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At least 631 records · Page 35Linked to original sources
PubMed2001

Use of whole genome sequence data to infer baculovirus phylogeny.

Several phylogenetic methods based on whole genome sequence data were evaluated using data from nine complete baculovirus genomes. The utility of three independent character sets was assessed. The first data set comprised the sequences of the 63 genes common to these viruses. The second set of characters was based on gene order, and phylogenies were inferred using both breakpoint distance analysis and a novel method developed here, termed neighbor pair analysis. The third set recorded gene content by scoring gene presence or absence in each genome. All three data sets yielded phylogenies supporting the separation of the Nucleopolyhedrovirus (NPV) and Granulovirus (GV) genera, the division of the NPVs into groups I and II, and species relationships within group I NPVs. Generation of phylogenies based on the combined sequences of all 63 shared genes proved to be the most effective approach to resolving the relationships among the group II NPVs and the GVs. The history of gene acquisitions and losses that have accompanied baculovirus diversification was visualized by mapping the gene content data onto the phylogenetic tree. This analysis highlighted the fluid nature of baculovirus genomes, with evidence of frequent genome rearrangements and multiple gene content changes during their evolution. Of more than 416 genes identified in the genomes analyzed, only 63 are present in all nine genomes, and 200 genes are found only in a single genome. Despite this fluidity, the whole genome-based methods we describe are sufficiently powerful to recover the underlying phylogeny of the viruses.

Animals↗
PubMed2026

Applicability of Nanopore-only whole-genome sequencing for Pseudomonas aeruginosa outbreak investigation in the ICU setting: a multicentric study.

UNLABELLED: Pseudomonas aeruginosa outbreaks frequently occur in intensive care units (ICUs). In particular, ICU patients requiring mechanical ventilation are vulnerable to P. aeruginosa ventilator-associated pneumonia, which is associated with high morbidity and mortality. Fast and accurate genotyping during the early stage is crucial to document and manage P. aeruginosa outbreaks at the ICU. In this study, we have evaluated the applicability of Oxford Nanopore whole-genome sequencing (WGS) for outbreak investigation and antimicrobial resistance (AMR) prediction. To evaluate whether a Nanopore-only WGS workflow was able to reproduce Illumina-confirmed transmission clusters, 19 P. aeruginosa isolates from ICUs at UZ Brussels (Belgium) that were previously sequenced with Illumina were sequenced using a Nanopore-only workflow based on the latest V14 chemistry, followed by bioinformatic analysis via BugSeq and MBioSEQ Ridom Typer. Although both bioinformatic platforms showed high concordance between Illumina and Nanopore data, MBioSEQ Ridom Typer yielded the lowest allelic distance (maximum one cgMLST allele), confirming all outbreak clusters. When applying the Nanopore-only workflow to longitudinally collected isolates, low genetic heterogeneity (maximum three cgMLST alleles) was observed between isolates from the same patient. WGS and subsequent outbreak analysis of 65 respiratory P. aeruginosa isolates collected from 38 different ICU patients across six Belgian hospitals during a 9-month period showed no intra- or inter-hospital transmission. When the Nanopore-only WGS data were used to predict AMR, there was high categorical agreement (95%) between AMR genotype and phenotype. These findings highlight the potential of Nanopore WGS as a rapid and accurate tool for outbreak investigation of P. aeruginosa. IMPORTANCE: In recent years, Nanopore sequencing has found its way to clinical laboratories because of its affordability, scalability, and, most importantly, its ability to obtain sequencing results in near-real time. However, despite improved raw read accuracies with the latest generation R10.4.1 flow cells, the question remains whether the achieved accuracy is sufficient for accurate bacterial outbreak investigation, particularly in high-risk settings such as intensive care units (ICUs). In this study, we show that Nanopore-only whole-genome sequencing (WGS) is able to match Illumina-only WGS in terms of accuracy for Pseudomonas aeruginosa outbreak investigation in the ICU setting, although important sequence type-dependent and even strain-specific methylation issues need to be resolved in order to guarantee this accuracy. By providing a fast and accurate workflow for reliable P. aeruginosa outbreak investigation, this study could pave the way for large-scale implementation of Nanopore-only WGS, leading to faster outbreak response times.

Humans↗
PubMed2004

Bovine enterovirus 2: complete genomic sequence and molecular modelling of a reference strain and a wild-type isolate from endemically infected US cattle.

Bovine enteroviruses are members of the family Picornaviridae, genus Enterovirus. Whilst little is known about their pathogenic potential, they are apparently endemic in some cattle and cattle environments. Only one of the two current serotypes has been sequenced completely. In this report, the entire genome sequences of bovine enterovirus 2 (BEV-2) strain PS87 and a recent isolate from an endemically infected herd in Maryland, USA (Wye3A) are presented. The recent isolate clearly segregated phylogenetically with sequences representing the BEV-2 serotype, as did other isolates from the endemic herd. The Wye3A isolate shared 82 % nucleotide sequence identity with the PS87 strain and 68 % identity with a BEV-1 strain (VG5-27). Comparison of BEV-2 and BEV-1 deduced protein sequences revealed 72-73 % identity and showed that most differences were single amino acid changes or single deletions, with the exception of the VP1 protein, where both BEV-2 sequences were 7 aa shorter than that of BEV-1. Homology modelling of the capsid proteins of BEV-2 against protein database entries for picornaviruses indicated six significant differences among bovine enteroviruses and other members of the family Picornaviridae. Five of these were on the 'rim' of the proposed enterovirus receptor-binding site or 'canyon' (VP1) and one was near the base of the canyon (VP3). Two of these regions varied enough to distinguish BEV-2 from BEV-1 strains. This is the first report and analysis of full-length sequences for BEV-2. Continued analysis of these wild-type strains should yield useful information for genotyping enteroviruses and modelling enterovirus capsid structure.

Animals↗
PubMed2002

Genome sequence of Shigella flexneri 2a: insights into pathogenicity through comparison with genomes of Escherichia coli K12 and O157.

We have sequenced the genome of Shigella flexneri serotype 2a, the most prevalent species and serotype that causes bacillary dysentery or shigellosis in man. The whole genome is composed of a 4 607 203 bp chromosome and a 221 618 bp virulence plasmid, designated pCP301. While the plasmid shows minor divergence from that sequenced in serotype 5a, striking characteristics of the chromosome have been revealed. The S.flexneri chromosome has, astonishingly, 314 IS elements, more than 7-fold over those possessed by its close relatives, the non-pathogenic K12 strain and enterohemorrhagic O157:H7 strain of Escherichia coli. There are 13 translocations and inversions compared with the E.coli sequences, all involve a segment larger than 5 kb, and most are associated with deletions or acquired DNA sequences, of which several are likely to be bacteriophage-transmitted pathogenicity islands. Furthermore, S.flexneri, resembling another human-restricted enteric pathogen, Salmonella typhi, also has hundreds of pseudogenes compared with the E.coli strains. All of these could be subjected to investigations towards novel preventative and treatment strategies against shigellosis.

Amino Acid Sequence↗
PubMed2006

Comparison of Salmonella enterica serovar Typhimurium LT2 and non-LT2 salmonella genomic sequences, and genotyping of salmonellae by using PCR.

Genes of Salmonella enterica serovar Typhimurium LT2 expected to be specifically present in Salmonella were selected using the Basic Local Alignment Search Tool (BLAST) program. The 152 selected genes were compared with 11 genomic sequences of Salmonella serovars, including Salmonella enterica subsp. I and IIIb and Salmonella bongori (V), and were clustered into 17 groups by their comparison patterns. A total of 38 primer pairs were constructed to represent each of the 17 groups, and PCR was performed with various Salmonella subspecies including Salmonella enterica subsp. I, II, IIIa, IIIb, IV, VI, and V to evaluate a comprehensive DNA-based scheme for identification of Salmonella subspecies and the major disease-causing Salmonella serovars. Analysis of PCR results showed that Salmonella enterica subsp. I was critically divided from other subspecies, and Salmonella strains belonging to S. enterica subsp. I were clustered based on their serovars. In addition, genotypic relationships within S. enterica subsp. I by PCR results were investigated. Also, Salmonella signature genes, Salmonella enterica serovar Typhimurium signature genes, and Salmonella enterica subsp. I signature genes were demonstrated based on their PCR results. The described PCR method suggests a rapid and convenient method for identification of Salmonella serovars that can be used by nonspecialized laboratories. Genome sequence comparison can be a useful tool in epidemiologic and taxonomic studies of Salmonella.

Bacterial Typing Techniques↗
PubMed2005

Partial mitochondrial genome sequences of Ostrinia nubilalis and Ostrinia furnicalis.

Contiguous 14,535 and 14,536 nt near complete mitochondrial genome sequences respectively were obtained for Ostrinia nubilalis and Ostrinia furnicalis. Mitochondrial gene order was identical to that observed from Bombyx. Sequences comparatively showed 186 substitutions (1.3% sequence divergence), 170 CDS substitutions (131 at 3(rd) codon positions), and an excess of transition mutation likely resulting by purifying selection (d(N)/d(S) = omega congruent with 0.15). Overall substitution rates were significantly higher at 4-fold (5.2%) compared to 2-fold degenerate codons (2.6%). These are the 3(rd) and 4(th) lepidopteran mitochondrial genome reference sequences in GenBank and useful for comparative mitochondrial studies.

Animals↗
PubMed2003

Comparative full-length genome sequence analysis of 14 SARS coronavirus isolates and common mutations associated with putative origins of infection.

BACKGROUND: The cause of severe acute respiratory syndrome (SARS) has been identified as a new coronavirus. Whole genome sequence analysis of various isolates might provide an indication of potential strain differences of this new virus. Moreover, mutation analysis will help to develop effective vaccines. METHODS: We sequenced the entire SARS viral genome of cultured isolates from the index case (SIN2500) presenting in Singapore, from three primary contacts (SIN2774, SIN2748, and SIN2677), and one secondary contact (SIN2679). These sequences were compared with the isolates from Canada (TOR2), Hong Kong (CUHK-W1 and HKU39849), Hanoi (URBANI), Guangzhou (GZ01), and Beijing (BJ01, BJ02, BJ03, BJ04). FINDINGS: We identified 129 sequence variations among the 14 isolates, with 16 recurrent variant sequences. Common variant sequences at four loci define two distinct genotypes of the SARS virus. One genotype was linked with infections originating in Hotel M in Hong Kong, the second contained isolates from Hong Kong, Guangzhou, and Beijing with no association with Hotel M (p<0.0001). Moreover, other common sequence variants further distinguished the geographical origins of the isolates, especially between Singapore and Beijing. INTERPRETATION: Despite the recent onset of the SARS epidemic, genetic signatures are emerging that partition the worldwide SARS viral isolates into groups on the basis of contact source history and geography. These signatures can be used to trace sources of infection. In addition, a common variant associated with a non-conservative aminoacid change in the S1 region of the spike protein, suggests that immunological pressures might be starting to influence the evolution of the SARS virus in human populations.

Amino Acid Sequence↗
PubMed2026

Development of a low-coverage whole genome sequencing screen for apomixis using a diverse set of Malus germplasm.

In the past decade, plant biologists have made several major discoveries pertaining to the genetic basis of apomixis (clonal propagation by seed) that have shown promise in preserving high-value hybrid rice and sorghum genotypes. This progress was made possible by foundational gene discovery efforts in model species and natural apomicts, but pleiotropic obstacles still limit its broad agricultural adoption, especially in eudicots. Thus, it follows that investigations of novel apomicts should lead to the development of new molecular tools for plant breeding. The two most common ways to identify clonal seed production are flow-cytometry seed screens and genome sequencing to compare the DNA sequences of the maternal parent and progeny, traditionally using low-throughput markers. While flow-cytometry has been the dominant method for more than two decades, it provides indirect information on the genetics of a resulting embryo and can be ineffective in certain species. Here we developed a method using short-read whole-genome sequencing at moderately low coverage (averaging 3X and 6X) to screen diverse Malus genotypes maintained in a USDA germplasm collection for clonal seed production. In total, we sequenced 55 genotypes, 1,216 of their embryos, and identified 17 previously undescribed apomictic genotypes. Several more were detected with the flow cytometry seed screen, which helped resolve certain types of reproduction and sources of noise in low-coverage datasets. This low-pass screening-by-sequencing method is a relatively low-cost, rapid method for detecting apomictic genotypes in diverse plant germplasm and when used thoughtfully in conjunction with flow cytometry, provides a new way to visualize the genetic outcomes of sexual and asexual reproduction in plants.

Apomixis↗
PubMed2006

Genome sequencing and identification of gene function in rice.

Rice is known to be one of the most important crops for human consumption. As the model cereal crop, large-scale sequencing of rice genome must play quite important roles both in theoretical research and practical application in rice breeding, which announces the opening of another new way to resolve the world food crisis. At present, the emphasis of rice genome research has been transferred from structure genomics to functional analysis. The discovery of new genes and annotation of gene function was believed to be an important issue in functional genomics research. In this article, the sequencing and functional research of the rice genome were reviewed. These results may provide some useful clues for rice genetic engineering and breeding practices.

Computational Biology↗
PubMed1997

Human synaptotagmin V (SYT5): sequence, genomic structure, and chromosomal location.

We have determined the sequence, genomic structure, and chromosomal location of the human synaptotagmin V (SYTV) gene. The human SYTV gene encodes a 386-amino-acid product which is 91% identical to rat Syt V. The human SYTV open reading frame is interrupted by seven introns which can be alternatively spliced. Human SYTV was found to lie very close to SYTIII on chromosome 19q13.4 by PCR analysis of somatic cell hybrid DNA and by DNA hybridization to arrayed cosmids of the chromosome 19 metric physical map. This provides the first report of linked synaptotagmin genes.

Amino Acid Sequence↗
PubMed2003

Efficiency improvement of peptide identification for an organism without complete genome sequence, using expressed sequence tag database and tandem mass spectral data.

We compared peptide identification by database (DB) search methods with de novo sequencing results for proteomics study in an organism without genome sequence information. When the former was done by searching the Expressed Sequence Tag (EST) DB of the sample organism or the NCBI nonredundant (nr) protein DB of green plants using either the MASCOT or SEQUEST software program, it was confirmed that the former is as accurate as the latter. Peptides identified from EST DB were twice as many as those from the nr protein DB, in spite of the fact that the EST DB has less data (26 222 EST) than the NCBI nr protein DB (224 238). This study demonstrates that EST DB with tandem mass spectra can be used reliably for high-throughput proteomics studies in an organism without genome information.

Algorithms↗
PubMed1993

Genome sequencing on both strands: the Janus strategy.

The design of large scale DNA sequencing projects such as genome analysis demands a new approach to sequencing strategy, since neither a purely random nor a purely directed method is satisfactory. We have developed a strategy that combines these two methods in a way that preserves the advantages of both while avoiding their particular limitations. Computer simulations showed that a specific balance of random and directed sequencing was required for the most efficient strategy, termed the Janus strategy, which has been used in the Escherichia coli genome sequencing project. This approach depended on obtaining sequence easily from either strand of a cloned insert, and was facilitated by inversion of the insert in the engineered M13 vector Janus, by site-specific recombination. The inversion was accomplished simply by growth on the appropriate host strain, when the DNA strand incorporated into the new single stranded phage was complementary to that in the original phage, and was sequenced by the same simple protocol as the first strand.

Bacteriophage lambda↗
PubMed2001

Complete genome sequence of a virulent isolate of Streptococcus pneumoniae.

The 2,160,837-base pair genome sequence of an isolate of Streptococcus pneumoniae, a Gram-positive pathogen that causes pneumonia, bacteremia, meningitis, and otitis media, contains 2236 predicted coding regions; of these, 1440 (64%) were assigned a biological role. Approximately 5% of the genome is composed of insertion sequences that may contribute to genome rearrangements through uptake of foreign DNA. Extracellular enzyme systems for the metabolism of polysaccharides and hexosamines provide a substantial source of carbon and nitrogen for S. pneumoniae and also damage host tissues and facilitate colonization. A motif identified within the signal peptide of proteins is potentially involved in targeting these proteins to the cell surface of low-guanine/cytosine (GC) Gram-positive species. Several surface-exposed proteins that may serve as potential vaccine candidates were identified. Comparative genome hybridization with DNA arrays revealed strain differences in S. pneumoniae that could contribute to differences in virulence and antigenicity.

Antigens, Bacterial↗
PubMed2004

Isolation, molecular characterisation and genome sequence of a bacteriophage (Chp3) from Chlamydophila pecorum.

Chlamydiae are obligate intracellular pathogens that have a unique developmental cycle. Thirty nine viable isolates representing all nine currently recognised chlamydial species were screened by immunofluorescence with a cross-reacting chlamydiaphage monoclonal antibody. A novel chlamydiaphage (Chp3) was detected in C. pecorum, a chlamydial species not previously known to carry bacteriophages. Chp3 belongs to the Microviridae, members of this virus family are characterised by circular, single-stranded DNA genomes and small T = 1 icosahedral capsids. Double-stranded replicative form Chp3 DNA was purified from elementary bodies and used as a template to determine the complete genome sequence. The genome of Chp3 is 4,554 base pairs and encodes eight open reading frames organised in the same genome structure as other chlamydiaphages. An unrooted phylogenetic tree was constructed based on the major coat proteins of 11 members of the Microviridae and Chp3. This showed that the Microviridae are clearly divided into two discrete sub-families; those that infect the Enterobacteriaceae e.g. ØX174 and the bacteriophages that infect obligate intracellular bacteria or mollicutes including SpV4 (Spiroplasma melliferum), ØMH2K (Bdellovibrio bacteriovorus) and the chlamydiaphages. Comparative analyses demonstrate that the chlamydiaphages can be further subdivided into two groupings, one represented by Chp2/Chp3 and the other by ØCPG1/ØCPAR39.

Base Sequence↗
PubMed2005

Evaluation of regulatory potential and conservation scores for detecting cis-regulatory modules in aligned mammalian genome sequences.

Techniques of comparative genomics are being used to identify candidate functional DNA sequences, and objective evaluations are needed to assess their effectiveness. Different analytical methods score distinctive features of whole-genome alignments among human, mouse, and rat to predict functional regions. We evaluated three of these methods for their ability to identify the positions of known regulatory regions in the well-studied HBB gene complex. Two methods, multispecies conserved sequences and phastCons, quantify levels of conservation to estimate a likelihood that aligned DNA sequences are under purifying selection. A third function, regulatory potential (RP), measures the similarity of patterns in the alignments to those in known regulatory regions. The methods can correctly identify 50%-60% of noncoding positions in the HBB gene complex as regulatory or nonregulatory, with RP performing better than do other methods. When evaluated by the ability to discriminate genomic intervals, RP reaches a sensitivity of 0.78 and a true discovery rate of approximately 0.6. The performance is better on other reference sets; both phastCons and RP scores can capture almost all regulatory elements in those sets along with approximately 7% of the human genome.

Algorithms↗