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1.
Viruses ; 8(12)2016 12 10.
Article in English | MEDLINE | ID: mdl-27973399

ABSTRACT

Vaccinia virus (VACV) has been implicated in infections of dairy cattle and humans, and outbreaks have substantially impacted local economies and public health in Brazil. During a 2005 outbreak, a VACV strain designated Serro 2 virus (S2V) was collected from a 30-year old male milker. Our aim was to phenotypically and genetically characterize this VACV Brazilian isolate. S2V produced small round plaques without associated comets when grown in BSC40 cells. Furthermore, S2V was less virulent than the prototype strain VACV-Western Reserve (WR) in a murine model of intradermal infection, producing a tiny lesion with virtually no surrounding inflammation. The genome of S2V was sequenced by primer walking. The coding region spans 184,572 bp and contains 211 predicted genes. Mutations in envelope genes specifically associated with small plaque phenotypes were not found in S2V; however, other alterations in amino acid sequences within these genes were identified. In addition, some immunomodulatory genes were truncated in S2V. Phylogenetic analysis using immune regulatory-related genes, besides the hemagglutinin gene, segregated the Brazilian viruses into two clusters, grouping the S2V into Brazilian VACV group 1. S2V is the first naturally-circulating human-associated VACV, with a low passage history, to be extensively genetically and phenotypically characterized.


Subject(s)
Genome, Viral , Phylogeny , Sequence Analysis, DNA , Vaccinia virus/genetics , Vaccinia virus/isolation & purification , Vaccinia/virology , Adult , Animals , Brazil , Cell Line , Disease Models, Animal , Genes, Viral , Humans , Male , Mice , Sequence Homology , Vaccinia/pathology , Vaccinia virus/classification , Vaccinia virus/physiology , Viral Plaque Assay , Virulence , Virulence Factors/genetics
2.
Emerg Infect Dis ; 17(11): 2113-21, 2011 Nov.
Article in English | MEDLINE | ID: mdl-22099115

ABSTRACT

Cholera was absent from the island of Hispaniola at least a century before an outbreak that began in Haiti in the fall of 2010. Pulsed-field gel electrophoresis (PFGE) analysis of clinical isolates from the Haiti outbreak and recent global travelers returning to the United States showed indistinguishable PFGE fingerprints. To better explore the genetic ancestry of the Haiti outbreak strain, we acquired 23 whole-genome Vibrio cholerae sequences: 9 isolates obtained in Haiti or the Dominican Republic, 12 PFGE pattern-matched isolates linked to Asia or Africa, and 2 nonmatched outliers from the Western Hemisphere. Phylogenies for whole-genome sequences and core genome single-nucleotide polymorphisms showed that the Haiti outbreak strain is genetically related to strains originating in India and Cameroon. However, because no identical genetic match was found among sequenced contemporary isolates, a definitive genetic origin for the outbreak in Haiti remains speculative.


Subject(s)
Genome, Bacterial , Vibrio cholerae/genetics , Africa/epidemiology , Alleles , Asia/epidemiology , Bacterial Proteins/genetics , Cholera/epidemiology , Cholera Toxin/genetics , Disease Outbreaks , Electrophoresis, Gel, Pulsed-Field , Gene Order , Haiti/epidemiology , Humans , Interspersed Repetitive Sequences/genetics , Phylogeny , Prophages , Sequence Homology, Amino Acid , Vibrio cholerae/classification , Vibrio cholerae/isolation & purification
3.
Database (Oxford) ; 2011: bar035, 2011.
Article in English | MEDLINE | ID: mdl-21930505

ABSTRACT

Neisseria meningitidis is an important pathogen, causing life-threatening diseases including meningitis, septicemia and in some cases pneumonia. Genomic studies hold great promise for N. meningitidis research, but substantial database resources are needed to deal with the wealth of information that comes with completely sequenced and annotated genomes. To address this need, we developed Neisseria Base (NBase), a comparative genomics database and genome browser that houses and displays publicly available N. meningitidis genomes. In addition to existing N. meningitidis genome sequences, we sequenced and annotated 19 new genomes using 454 pyrosequencing and the CG-Pipeline genome analysis tool. In total, NBase hosts 27 complete N. meningitidis genome sequences along with their associated annotations. The NBase platform is designed to be scalable, via the underlying database schema and modular code architecture, such that it can readily incorporate new genomes and their associated annotations. The front page of NBase provides user access to these genomes through searching, browsing and downloading. NBase search utility includes BLAST-based sequence similarity searches along with a variety of semantic search options. All genomes can be browsed using a modified version of the GBrowse platform, and a plethora of information on each gene can be viewed using a customized details page. NBase also has a whole-genome comparison tool that yields single-nucleotide polymorphism differences between two user-defined groups of genomes. Using the virulent ST-11 lineage as an example, we demonstrate how this comparative genomics utility can be used to identify novel genomic markers for molecular profiling of N. meningitidis.


Subject(s)
Database Management Systems , Genome, Bacterial , Genomics/methods , Neisseria meningitidis/genetics , Sequence Analysis, DNA/methods , Genetic Markers , Polymorphism, Single Nucleotide , User-Computer Interface
4.
PLoS One ; 6(8): e23086, 2011.
Article in English | MEDLINE | ID: mdl-21858000

ABSTRACT

Cowpox virus (CPXV) is described as the source of the first vaccine used to prevent the onset and spread of an infectious disease. It is one of the earliest described members of the genus Orthopoxvirus, which includes the viruses that cause smallpox and monkeypox in humans. Both the historic and current literature describe "cowpox" as a disease with a single etiologic agent. Genotypic data presented herein indicate that CPXV is not a single species, but a composite of several (up to 5) species that can infect cows, humans, and other animals. The practice of naming agents after the host in which the resultant disease manifests obfuscates the true taxonomic relationships of "cowpox" isolates. These data support the elevation of as many as four new species within the traditional "cowpox" group and suggest that both wild and modern vaccine strains of Vaccinia virus are most closely related to CPXV of continental Europe rather than the United Kingdom, the homeland of the vaccine.


Subject(s)
Cowpox virus/classification , Cowpox virus/genetics , Genome, Viral/genetics , Phylogeny , Animals , Cluster Analysis , Cowpox virus/isolation & purification , DNA, Viral/chemistry , DNA, Viral/genetics , Humans , Molecular Sequence Data , Sequence Analysis, DNA , Species Specificity , Viral Proteins/genetics
5.
Antimicrob Agents Chemother ; 54(10): 4314-20, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20660665

ABSTRACT

Of the 9 vancomycin-resistant Staphylococcus aureus (VRSA) cases reported to date in the literature, 7 occurred in Michigan. In 5 of the 7 Michigan VRSA cases, an Inc18-like vanA plasmid was identified in the VRSA isolate and/or an associated vancomycin-resistant Enterococcus (VRE) isolate from the same patient. This plasmid may play a critical role in the emergence of VRSA. We studied the geographical distribution of the plasmid by testing 1,641 VRE isolates from three separate collections by PCR for plasmid-specific genes traA, repR, and vanA. Isolates from one collection (phase 2) were recovered from surveillance cultures collected in 17 hospitals in 13 states. All VRE isolates from 2 Michigan institutions (n = 386) and between 60 and 70 VRE isolates (n = 883) from the other hospitals were tested. Fifteen VRE isolates (3.9%) from Michigan were positive for an Inc18-like vanA plasmid (9 E. faecalis [12.5%], 3 E. faecium [1.0%], 2 E. avium, and 1 E. raffinosus). Six VRE isolates (0.6%) from outside Michigan were positive (3 E. faecalis [2.7%] and 3 E. faecium [0.4%]). Of all E. faecalis isolates tested, 6.0% were positive for the plasmid, compared to 0.6% for E. faecium and 3.0% for other spp. Fourteen of the 15 plasmid-positive isolates from Michigan had the same Tn1546 insertion site location as the VRSA-associated Inc18-like plasmid, whereas 5 of 6 plasmid-positive isolates from outside Michigan differed in this characteristic. Most plasmid-positive E. faecalis isolates demonstrated diverse patterns by PFGE, with the exception of three pairs with indistinguishable patterns, suggesting that the plasmid is mobile in nature. Although VRE isolates with the VRSA-associated Inc18-like vanA plasmid were more common in Michigan, they remain rare. Periodic surveillance of VRE isolates for the plasmid may be useful in predicting the occurrence of VRSA.


Subject(s)
Enterococcus/drug effects , Enterococcus/genetics , Plasmids/genetics , Staphylococcus aureus/genetics , Vancomycin Resistance/genetics , Bacterial Proteins/genetics , Electrophoresis, Gel, Pulsed-Field , Enterococcus faecalis/drug effects , Enterococcus faecalis/genetics , Enterococcus faecium/drug effects , Enterococcus faecium/genetics , Polymerase Chain Reaction , Staphylococcus aureus/drug effects
6.
J Bacteriol ; 192(4): 1030-44, 2010 Feb.
Article in English | MEDLINE | ID: mdl-20008069

ABSTRACT

Legionella longbeachae causes most cases of legionellosis in Australia and may be underreported worldwide due to the lack of L. longbeachae-specific diagnostic tests. L. longbeachae displays distinctive differences in intracellular trafficking, caspase 1 activation, and infection in mouse models compared to Legionella pneumophila, yet these two species have indistinguishable clinical presentations in humans. Unlike other legionellae, which inhabit freshwater systems, L. longbeachae is found predominantly in moist soil. In this study, we sequenced and annotated the genome of an L. longbeachae clinical isolate from Oregon, isolate D-4968, and compared it to the previously published genomes of L. pneumophila. The results revealed that the D-4968 genome is larger than the L. pneumophila genome and has a gene order that is different from that of the L. pneumophila genome. Genes encoding structural components of type II, type IV Lvh, and type IV Icm/Dot secretion systems are conserved. In contrast, only 42/140 homologs of genes encoding L. pneumophila Icm/Dot substrates have been found in the D-4968 genome. L. longbeachae encodes numerous proteins with eukaryotic motifs and eukaryote-like proteins unique to this species, including 16 ankyrin repeat-containing proteins and a novel U-box protein. We predict that these proteins are secreted by the L. longbeachae Icm/Dot secretion system. In contrast to the L. pneumophila genome, the L. longbeachae D-4968 genome does not contain flagellar biosynthesis genes, yet it contains a chemotaxis operon. The lack of a flagellum explains the failure of L. longbeachae to activate caspase 1 and trigger pyroptosis in murine macrophages. These unique features of L. longbeachae may reflect adaptation of this species to life in soil.


Subject(s)
Bacterial Proteins/genetics , Genome, Bacterial , Legionella longbeachae/genetics , Legionella longbeachae/pathogenicity , Legionellosis/microbiology , Sequence Analysis, DNA , Virulence Factors/genetics , Aged , Conserved Sequence , Female , Humans , Legionella longbeachae/isolation & purification , Legionella pneumophila/genetics , Molecular Sequence Data , Oregon , Synteny
7.
PLoS One ; 4(10): e7666, 2009 Oct 29.
Article in English | MEDLINE | ID: mdl-19865479

ABSTRACT

The data presented herein support the North American orthopoxviruses (NA OPXV) in a sister relationship to all other currently described Orthopoxvirus (OPXV) species. This phylogenetic analysis reaffirms the identification of the NA OPXV as close relatives of "Old World" (Eurasian and African) OPXV and presents high support for deeper nodes within the Chordopoxvirinae family. The natural reservoir host(s) for many of the described OPXV species remains unknown although a clear virus-host association exists between the genus OPXV and several mammalian taxa. The hypothesized host associations and the deep divergence of the OPXV/NA OPXV clades depicted in this study may reflect the divergence patterns of the mammalian faunas of the Old and New World and reflect a more ancient presence of OPXV on what are now the American continents. Genes from the central region of the poxvirus genome are generally more conserved than genes from either end of the linear genome due to functional constraints imposed on viral replication abilities. The relatively slower evolution of these genes may more accurately reflect the deeper history among the poxvirus group, allowing for robust placement of the NA OPXV within Chordopoxvirinae. Sequence data for nine genes were compiled from three NA OPXV strains plus an additional 50 genomes collected from Genbank. The current, gene sequence based phylogenetic analysis reaffirms the identification of the NA OPXV as the nearest relatives of "Old World" OPXV and presents high support for deeper nodes within the Chordopoxvirinae family. Additionally, the substantial genetic distances that separate the currently described NA OPXV species indicate that it is likely that many more undescribed OPXV/NA OPXV species may be circulating among wild animals in North America.


Subject(s)
Orthopoxvirus/classification , Orthopoxvirus/genetics , DNA, Mitochondrial/genetics , DNA, Viral/genetics , Databases, Factual , Ecology , Ecosystem , Evolution, Molecular , Genetic Speciation , Geography , North America , Phylogeny , Sequence Analysis, DNA , Species Specificity
8.
Vaccine ; 25(52): 8807-32, 2007 Dec 17.
Article in English | MEDLINE | ID: mdl-18037545

ABSTRACT

Conventional vaccines used for smallpox eradication were often denoted one or another strain of Vaccinia virus (VACV), even though seed virus was sub-cultured multifariously, which rendered the virion population genetically heterogeneous. ACAM2000 cell culture vaccine, recently licensed in the U.S., consists of a biologically vaccine-like VACV homogeneous-sequence clone from the conventional smallpox vaccine Dryvax, which we verified from Dryvax sequence chromatograms is genetically heterogeneous. ACAM2000 VACV and CL3, a mouse-neurovirulent clone from Dryvax, differ by 572 single nucleotide polymorphisms and 53 insertions-deletions of varied size, including a 4.5-kbp deletion in ACAM2000 and a 6.2-kbp deletion in CL3. The sequence diversity between the two clones precludes precisely defining why CL3 is more pathogenic; however, four genes appear significantly dissimilar to account for virulence differences. CL3 encodes intact immunomodulators interferon-alpha/beta and tumor necrosis factor receptors, which are truncated in ACAM2000. CL3 specifies a Cowpox and Variola virus-like ankyrin-repeat protein that might be associated with proteolysis via ubiquitination. And, CL3 shows an elongated thymidylate kinase, similar to the enzyme of the mouse-neurovirulent VACV-WR, a derivative of the New York City Board of Health vaccine, the origin vaccine of Dryvax. Although ACAM2000 encodes most proteins associated with immunization protection, the cloning probably delimited the variant epitopes and other motifs produced by Dryvax due to its VACV genetic heterogeneity. The sequence information for ACAM2000 and CL3 could be significant for resolving the dynamics of their different proteomes and thereby aid development of safer, more effective vaccines.


Subject(s)
Genome, Viral , Smallpox Vaccine/genetics , Vaccinia virus/genetics , Vaccinia virus/pathogenicity , Animals , Gene Order , Mice , Molecular Sequence Data , Point Mutation , Polymorphism, Genetic , Recombination, Genetic , Sequence Analysis, DNA , Sequence Deletion , Synteny , United States , Viral Proteins/genetics
9.
Virus Res ; 129(1): 11-25, 2007 Oct.
Article in English | MEDLINE | ID: mdl-17574698

ABSTRACT

Members of the genus Yatapoxvirus, which include Tanapox virus (TPV) and Yaba monkey tumor virus, infect primates including humans. Two strains of TPV isolated 50 years apart from patients infected from the equatorial region of Africa have been sequenced. The original isolate from a human case in the Tana River Valley, Kenya, in 1957 (TPV-Kenya) and an isolate from an infected traveler in the Republic of Congo in 2004 (TPV-RoC). Although isolated 50 years apart the genomes were highly conserved. The genomes differed at only 35 of 144,565 nucleotide positions (99.98% identical). We predict that TPV-RoC encodes 155 ORFs, however a single transversion (at nucleotide 10241) in TPV-Kenya resulted in the coding capacity for two predicted ORFs (11.1L and 11.2L) in comparison to a single ORF (11L) in TPV-RoC. The genomes of TPV are A+T rich (73%) and 96% of the sequence encodes predicted ORFs. Comparative genomic analysis identified several features shared with other chordopoxviruses. A conserved sequence within the terminal inverted repeat region that is also present in the other members of the Yatapoxviruses as well as members of the Capripoxviruses, Swinepox virus and an unclassified Deerpox virus suggests the existence of a conserved near-terminal sequence secondary structure. Two previously unidentified gene families were annotated that are represented by ORF TPV28L, which matched homologues in certain other chordopoxviruses, and TPV42.5L, which is highly conserved among currently reported chordopoxvirus sequences.


Subject(s)
DNA, Viral/genetics , Genome, Viral , Poxviridae Infections/virology , Tumor Virus Infections/virology , Yatapoxvirus/genetics , Africa , Amino Acid Sequence , Humans , Molecular Sequence Data , Open Reading Frames , Sequence Alignment , Sequence Homology
10.
Science ; 313(5788): 807-12, 2006 Aug 11.
Article in English | MEDLINE | ID: mdl-16873609

ABSTRACT

Comparative genomics of 45 epidemiologically varied variola virus isolates from the past 30 years of the smallpox era indicate low sequence diversity, suggesting that there is probably little difference in the isolates' functional gene content. Phylogenetic clustering inferred three clades coincident with their geographical origin and case-fatality rate; the latter implicated putative proteins that mediate viral virulence differences. Analysis of the viral linear DNA genome suggests that its evolution involved direct descent and DNA end-region recombination events. Knowing the sequences will help understand the viral proteome and improve diagnostic test precision, therapeutics, and systems for their assessment.


Subject(s)
DNA, Viral/genetics , Evolution, Molecular , Genetic Variation , Genome, Viral , Smallpox/virology , Variola virus/genetics , Disease Outbreaks , Gene Deletion , Genomics , Humans , Molecular Sequence Data , Open Reading Frames , Phylogeny , Proteome/analysis , Proteome/genetics , Recombination, Genetic , Sequence Analysis, DNA , Smallpox/epidemiology , Smallpox/mortality , Variola virus/classification , Variola virus/isolation & purification , Variola virus/pathogenicity , Viral Proteins/chemistry , Viral Proteins/genetics , Virulence/genetics
11.
Appl Environ Microbiol ; 72(1): 207-11, 2006 Jan.
Article in English | MEDLINE | ID: mdl-16391044

ABSTRACT

Severe acute respiratory syndrome (SARS) was discovered during a recent global outbreak of atypical pneumonia. A number of immunologic and molecular studies of the clinical samples led to the conclusion that a novel coronavirus (SARS-CoV) was associated with the outbreak. Later, a SARS resequencing GeneChip was developed by Affymetrix to characterize the complete genome of SARS-CoV on a single GeneChip. The present study was carried out to evaluate the performance of SARS resequencing GeneChips. Two human SARS-CoV strains (CDC#200301157 and Urbani) were resequenced by the SARS GeneChips. Five overlapping PCR amplicons were generated for each strain and hybridized with these GeneChips. The successfully hybridized GeneChips generated nucleotide sequences of nearly complete genomes for the two SARS-CoV strains with an average call rate of 94.6%. Multiple alignments of nucleotide sequences obtained from SARS GeneChips and conventional sequencing revealed full concordance. Furthermore, the GeneChip-based analysis revealed no additional polymorphic sites. The results of this study suggest that GeneChip-based genome characterization is fast and reproducible. Thus, SARS resequencing GeneChips may be employed as an alternate tool to obtain genome sequences of SARS-CoV strains pathogenic for humans in order to further understand the transmission dynamics of these viruses.


Subject(s)
Genome, Viral , Oligonucleotide Array Sequence Analysis , Sequence Analysis, DNA , Severe Acute Respiratory Syndrome/virology , Severe acute respiratory syndrome-related coronavirus/classification , Humans , Oligonucleotide Array Sequence Analysis/instrumentation , Reproducibility of Results , Severe acute respiratory syndrome-related coronavirus/genetics , Time Factors
12.
J Gen Virol ; 86(Pt 10): 2661-2672, 2005 Oct.
Article in English | MEDLINE | ID: mdl-16186219

ABSTRACT

Human monkeypox was first recognized outside Africa in 2003 during an outbreak in the USA that was traced to imported monkeypox virus (MPXV)-infected West African rodents. Unlike the smallpox-like disease described in the Democratic Republic of the Congo (DRC; a Congo Basin country), disease in the USA appeared milder. Here, analyses compared clinical, laboratory and epidemiological features of confirmed human monkeypox case-patients, using data from outbreaks in the USA and the Congo Basin, and the results suggested that human disease pathogenicity was associated with the viral strain. Genomic sequencing of USA, Western and Central African MPXV isolates confirmed the existence of two MPXV clades. A comparison of open reading frames between MPXV clades permitted prediction of viral proteins that could cause the observed differences in human pathogenicity between these two clades. Understanding the molecular pathogenesis and clinical and epidemiological properties of MPXV can improve monkeypox prevention and control.


Subject(s)
DNA, Viral/genetics , Genome, Viral , Monkeypox virus/classification , Phylogeny , Poxviridae Infections/epidemiology , Animals , Molecular Sequence Data , Monkeypox virus/genetics , Open Reading Frames , Poxviridae Infections/diagnosis , Sequence Homology, Amino Acid , Sequence Homology, Nucleic Acid
13.
Science ; 300(5624): 1394-9, 2003 May 30.
Article in English | MEDLINE | ID: mdl-12730500

ABSTRACT

In March 2003, a novel coronavirus (SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS). The sequence of the complete genome of SARS-CoV was determined, and the initial characterization of the viral genome is presented in this report. The genome of SARS-CoV is 29,727 nucleotides in length and has 11 open reading frames, and its genome organization is similar to that of other coronaviruses. Phylogenetic analyses and sequence comparisons showed that SARS-CoV is not closely related to any of the previously characterized coronaviruses.


Subject(s)
Genome, Viral , RNA, Viral/genetics , Severe acute respiratory syndrome-related coronavirus/genetics , Viral Proteins/genetics , Amino Acid Sequence , Conserved Sequence , Coronavirus/classification , Coronavirus/genetics , Coronavirus M Proteins , Coronavirus Nucleocapsid Proteins , DNA, Complementary , Endopeptidases/chemistry , Endopeptidases/genetics , Humans , Membrane Glycoproteins/chemistry , Membrane Glycoproteins/genetics , Molecular Sequence Data , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/genetics , Open Reading Frames , Phylogeny , Polyproteins/chemistry , Polyproteins/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA-Dependent RNA Polymerase/chemistry , RNA-Dependent RNA Polymerase/genetics , Regulatory Sequences, Nucleic Acid , Severe acute respiratory syndrome-related coronavirus/chemistry , Severe acute respiratory syndrome-related coronavirus/classification , Severe acute respiratory syndrome-related coronavirus/isolation & purification , Sequence Analysis, DNA , Severe Acute Respiratory Syndrome/virology , Spike Glycoprotein, Coronavirus , Transcription, Genetic , Viral Envelope Proteins/chemistry , Viral Envelope Proteins/genetics , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/genetics , Viral Proteins/chemistry
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