RESUMO
Continued advancements in sequencing technologies have fueled the development of new sequencing applications and promise to flood current databases with raw data. A number of factors prevent the seamless and easy use of these data, including the breadth of project goals, the wide array of tools that individually perform fractions of any given analysis, the large number of associated software/hardware dependencies, and the detailed expertise required to perform these analyses. To address these issues, we have developed an intuitive web-based environment with a wide assortment of integrated and cutting-edge bioinformatics tools in pre-configured workflows. These workflows, coupled with the ease of use of the environment, provide even novice next-generation sequencing users with the ability to perform many complex analyses with only a few mouse clicks and, within the context of the same environment, to visualize and further interrogate their results. This bioinformatics platform is an initial attempt at Empowering the Development of Genomics Expertise (EDGE) in a wide range of applications for microbial research.
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Bacillus anthracis/classificação , Biologia Computacional/métodos , Ebolavirus/classificação , Escherichia coli/classificação , Software , Yersinia pestis/classificação , Antraz/microbiologia , Bacillus anthracis/genética , Ebolavirus/genética , Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Doença pelo Vírus Ebola/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Internet , Filogenia , Peste/microbiologia , Yersinia pestis/genéticaRESUMO
The taxonomic status of the bacterium Wolbachia persica is described, and based on the evidence presented, transfer of this species to the genus Francisella as Francisella persica comb. nov. is proposed. This reclassification is supported by data generated from genomic comparisons of W. persica ATCC VR-331T ( = FSC845T = DSM 101678T) to other near neighbours, including Francisella tularensis subsp. novicida. The full-length 16S rRNA gene sequence of strain ATCC VR-331T had 98.5 % nucleotide identity to the cognate gene in F. tularensis, with the highest similarity to subspecies novicida. Phylogenetic trees of full-length 16S rRNA gene, gyrA and recA sequences from species of the genera Wolbachia (class Alphaproteobacteria) and Francisella (class Gammaproteobacteria) indicated that W. persica ATCC VR-331T was most closely related to members of the genus Francisella and not Wolbachia. Local collinear blocks within the chromosome of strain ATCC VR-331T had considerable similarity with F. tularensis subsp. novicida, but not with any Wolbachia strain. The genomes of strain ATCC VR-331T and F. tularensis subsp. novicida Utah 112T ( = ATCC 15482T) contained an average nucleotide identity mean of 88.72 % and median of 89.18 %. Importantly, the genome of strain ATCC VR-331T contained one Francisella Pathogenicity Island, similar to F. tularensis subsp. novicida, as well as the Francisella-specific gene fopA1 and F. tularensis-specific genes fopA2 and lpnA (also referred to as tul4). In contrast to the obligate intracellular genus Wolbachia, strain ATCC VR-331T and facultative intracellular Francisella can replicate in specialized cell-free media. Collectively, these results demonstrate that Wolbachia persica should be reclassified in the genus Francisella as Francisella persica comb. nov. The type strain of Francisella persica comb. nov. is ATCC VR-331T ( = FSC845T = DSM 101678T). An emended description of the family Francisellaceae is also provided.
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Francisella/classificação , Filogenia , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Wolbachia/classificaçãoRESUMO
Lack of trust in government-supported services after the death of a health care worker with symptoms of Ebola resulted in ongoing Ebola transmission in 2 Liberia counties. Ebola transmission was facilitated by attempts to avoid cremation of the deceased patient and delays in identifying and monitoring contacts.
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Doença pelo Vírus Ebola/diagnóstico , Adolescente , Adulto , Idoso , Análise por Conglomerados , Busca de Comunicante , Evolução Fatal , Feminino , Doença pelo Vírus Ebola/transmissão , Humanos , Libéria , MasculinoRESUMO
The henipaviruses, represented by Hendra (HeV) and Nipah (NiV) viruses are highly pathogenic zoonotic paramyxoviruses with uniquely broad host tropisms responsible for repeated outbreaks in Australia, Southeast Asia, India and Bangladesh. The high morbidity and mortality rates associated with infection and lack of licensed antiviral therapies make the henipaviruses a potential biological threat to humans and livestock. Henipavirus entry is initiated by the attachment of the G envelope glycoprotein to host cell membrane receptors. Previously, henipavirus-neutralizing human monoclonal antibodies (hmAb) have been isolated using the HeV-G glycoprotein and a human naïve antibody library. One cross-reactive and receptor-blocking hmAb (m102.4) was recently demonstrated to be an effective post-exposure therapy in two animal models of NiV and HeV infection, has been used in several people on a compassionate use basis, and is currently in development for use in humans. Here, we report the crystal structure of the complex of HeV-G with m102.3, an m102.4 derivative, and describe NiV and HeV escape mutants. This structure provides detailed insight into the mechanism of HeV and NiV neutralization by m102.4, and serves as a blueprint for further optimization of m102.4 as a therapeutic agent and for the development of entry inhibitors and vaccines.
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Anticorpos Monoclonais/química , Anticorpos Neutralizantes/química , Anticorpos Antivirais/química , Vírus Hendra/química , Proteínas Virais de Fusão/química , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Linhagem Celular , Reações Cruzadas , Cristalografia por Raios X , Vírus Hendra/genética , Vírus Hendra/imunologia , Infecções por Henipavirus/genética , Infecções por Henipavirus/imunologia , Humanos , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologiaRESUMO
In order to better characterize the Bacillus anthracis typing phage AP50c, we designed a genetic screen to identify its bacterial receptor. Insertions of the transposon mariner or targeted deletions of the structural gene for the S-layer protein Sap and the sporulation genes spo0A, spo0B, and spo0F in B. anthracis Sterne resulted in phage resistance with concomitant defects in phage adsorption and infectivity. Electron microscopy of bacteria incubated with AP50c revealed phage particles associated with the surface of bacilli of the Sterne strain but not with the surfaces of Δsap, Δspo0A, Δspo0B, or Δspo0F mutants. The amount of Sap in the S layer of each of the spo0 mutant strains was substantially reduced compared to that of the parent strain, and incubation of AP50c with purified recombinant Sap led to a substantial reduction in phage activity. Phylogenetic analysis based on whole-genome sequences of B. cereus sensu lato strains revealed several closely related B. cereus and B. thuringiensis strains that carry sap genes with very high similarities to the sap gene of B. anthracis. Complementation of the Δsap mutant in trans with the wild-type B. anthracis sap or the sap gene from either of two different B. cereus strains that are sensitive to AP50c infection restored phage sensitivity, and electron microscopy confirmed attachment of phage particles to the surface of each of the complemented strains. Based on these data, we postulate that Sap is involved in AP50c infectivity, most likely acting as the phage receptor, and that the spo0 genes may regulate synthesis of Sap and/or formation of the S layer.
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Fagos Bacilares/fisiologia , Bacillus anthracis/virologia , Proteínas de Bactérias/metabolismo , Interações Hospedeiro-Parasita , Ligação Viral , Fagos Bacilares/genética , Bacillus cereus/genética , Bacillus thuringiensis/genética , Proteínas de Bactérias/genética , Deleção de Genes , Teste de Complementação Genética , Mutagênese Insercional , Filogenia , Homologia de Sequência de AminoácidosRESUMO
BACKGROUND: The introduction of benchtop sequencers has made adoption of whole genome sequencing possible for a broader community of researchers than ever before. Concurrently, metagenomic sequencing (MGS) is rapidly emerging as a tool for interrogating complex samples that defy conventional analyses. In addition, next-generation sequencers are increasingly being used in clinical or related settings, for instance to track outbreaks. However, information regarding the analytical sensitivity or limit of detection (LoD) of benchtop sequencers is currently lacking. Furthermore, the specificity of sequence information at or near the LoD is unknown. RESULTS: In the present study, we assess the ability of three next-generation sequencing platforms to identify a pathogen (viral or bacterial) present in low titers in a clinically relevant sample (blood). Our results indicate that the Roche-454 Titanium platform is capable of detecting Dengue virus at titers as low as 1X102.5 pfu/mL, corresponding to an estimated 5.4X104 genome copies/ml maximum. The increased throughput of the benchtop sequencers, the Ion Torrent PGM and Illumina MiSeq platforms, enabled detection of viral genomes at concentrations as low as 1X104 genome copies/mL. Platform-specific biases were evident in sequence read distributions as well as viral genome coverage. For bacterial samples, only the MiSeq platform was able to provide sequencing reads that could be unambiguously classified as originating from Bacillus anthracis. CONCLUSION: The analytical sensitivity of all three platforms approaches that of standard qPCR assays. Although all platforms were able to detect pathogens at the levels tested, there were several noteworthy differences. The Roche-454 Titanium platform produced consistently longer reads, even when compared with the latest chemistry updates for the PGM platform. The MiSeq platform produced consistently greater depth and breadth of coverage, while the Ion Torrent was unequaled for speed of sequencing. None of the platforms were able to verify a single nucleotide polymorphism responsible for antiviral resistance in an Influenza A strain isolated from the 2009 H1N1 pandemic. Overall, the benchtop platforms perform well for identification of pathogens from a representative clinical sample. However, unlike identification, characterization of pathogens is likely to require higher titers, multiple libraries and/or multiple sequencing runs.
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Sequenciamento de Nucleotídeos em Larga Escala/instrumentação , Bacillus anthracis/genética , Mapeamento Cromossômico , Biologia Computacional , DNA Bacteriano/sangue , Bases de Dados Genéticas , Vírus da Dengue/genética , Genoma Viral , Sequenciamento de Nucleotídeos em Larga Escala/normas , Humanos , Vírus da Influenza A Subtipo H1N1/genética , RNA Viral/sangueRESUMO
BACKGROUND: Spontaneous Bacillus anthracis mutants resistant to infection by phage AP50c (AP50R) exhibit a mucoid colony phenotype and secrete an extracellular matrix. METHODS: Here we utilized a Roche/454-based whole genome sequencing approach to identify mutations that are candidates for conferring AP50c phage resistance, followed by genetic deletion and complementation studies to validate the whole genome sequence data and demonstrate that the implicated gene is necessary for AP50c phage infection. RESULTS: Using whole genome sequence data, we mapped the relevant mutations in six AP50R strains to csaB. Eleven additional spontaneous mutants, isolated in two different genetic backgrounds, were screened by PCR followed by Sanger sequencing of the csaB gene. In each spontaneous mutant, we found either a non-synonymous substitution, a nonsense mutation, or a frame-shift mutation caused by single nucleotide polymorphisms or a 5 base pair insertion in csaB. All together, 5 and 12 of the 17 spontaneous mutations are predicted to yield altered full length and truncated CsaB proteins respectively. As expected from these results, a targeted deletion or frame-shift mutations introduced into csaB in a different genetic background, in a strain not exposed to AP50c, resulted in a phage resistant phenotype. Also, substitution of a highly conserved histidine residue with an alanine residue (H270A) in CsaB resulted in phage resistance, suggesting that a functional CsaB is necessary for phage sensitivity. Conversely, introduction of the wild type allele of csaB in cis into the csaB deletion mutant by homologous recombination or supplying the wild type CsaB protein in trans from a plasmid restored phage sensitivity. The csaB mutants accumulated cell wall material and appeared to have a defective S-layer, whereas these phenotypes were reverted in the complemented strains. CONCLUSIONS: Taken together, these data suggest an essential role for csaB in AP50c phage infection, most likely in phage adsorption. (The whole genome sequences generated from this study have been submitted to GenBank under SRA project ID: SRA023659.1 and sample IDs: AP50 R1: SRS113675.1, AP50 R2: SRS113676.1, AP50 R3: SRS113728.1, AP50 R4: SRS113733.1, AP50 R6: SRS113734.1, JB220 Parent: SRS150209.1, JB220 Mutant: SRS150211.1).
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Fagos Bacilares/fisiologia , Bacillus anthracis/genética , Bacillus anthracis/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Genoma Bacteriano , Mutação , Sequência de Aminoácidos , Bacillus anthracis/ultraestrutura , Bacillus anthracis/virologia , Bacteriólise , Sequência de Bases , Mapeamento Cromossômico , Ordem dos Genes , Dados de Sequência Molecular , Óperon , Fenótipo , Plasmídeos/genética , Alinhamento de Sequência , Análise de Sequência de DNARESUMO
Respiratory symptoms are frequently reported in personnel deployed to the Middle East. This project characterized the respiratory toxicity of inhaled Iraqi sand (IS). Adult rats underwent a 6-wk inhalation to air or mainstream cigarette smoke (MSCS) (3 h/d, 5 d/wk) that included exposure to IS or crystalline silica (1 mg/m(3), 19 h/d, 7 d/wk) or air during the last 2 weeks. Assessments included motor activity, whole-body plethysmography, cytological and biochemical analysis of bronchoalveolar lavage fluid, lung metal burden, nasal and lung pathology, and changes in lung protein and gene expression. A number of metals including nickel, manganese, vanadium, and chromium were detected in IS. Elevated lung parenchyma aluminum, silica, barium, manganese, and vanadium concentrations were seen in IS-exposed rats, suggesting that several metals present in IS are bioavailable. Rats exposed to IS only developed mild inflammation in the anterior nose and lung. Silica inhalation was associated with some pulmonary responses that were not seen in IS-exposed rats, such as mild laryngeal and tracheal inflammation, mild tracheal epithelial hyperplasia, and elevated lung silica concentrations. MSCS inhalation with or without co-exposure to either IS or silica resulted in changes consistent with pulmonary inflammation and stress response. Rats exposed to MSCS and silica had more widespread airway lesions when compared with rats exposed to MSCS only. Silica-exposed rats had more robust pulmonary gene expression and proteomic responses than that seen in IS-exposed rat. Our studies show that the respiratory toxicity of IS is qualitatively similar to or less than that seen following short-term silica exposure.
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Poluentes Atmosféricos/toxicidade , Poeira , Metais/toxicidade , Dióxido de Silício/toxicidade , Poluição por Fumaça de Tabaco/efeitos adversos , Administração por Inalação , Animais , Comportamento Animal/efeitos dos fármacos , Testes de Provocação Brônquica , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/citologia , Poeira/análise , Expressão Gênica/efeitos dos fármacos , Força da Mão , Iraque , Laringe/efeitos dos fármacos , Laringe/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Pulmão/fisiologia , Masculino , Metais/análise , Atividade Motora/efeitos dos fármacos , Análise de Sequência com Séries de Oligonucleotídeos , Ratos , Ratos Sprague-Dawley , Testes de Função Respiratória , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/patologia , Dióxido de Silício/análise , Traqueia/efeitos dos fármacos , Traqueia/patologiaRESUMO
Heavy metal tungsten alloys have replaced lead and depleted uranium in many munitions applications, due to public perception of these elements as environmentally unsafe. Tungsten materials left in the environment may become bioaccessible as tungstate, which might lead to population exposure through water and soil contamination. Although tungsten had been considered a relatively inert and toxicologically safe material, recent research findings have raised concerns about possible deleterious health effects after acute and chronic exposure to this metal. This investigation describes tissue distribution of tungsten in mice following oral exposure to sodium tungstate. Twenty-four 6-9 weeks-old C57BL/6 laboratory mice were exposed to different oral doses of sodium tungstate (0, 62.5, 125, and 200 mg/kg/d) for 28 days, and after one day, six organs were harvested for trace element analysis with inductively coupled plasma mass spectrometry (ICP-MS). Kidney, liver, colon, bone, brain, and spleen were analyzed by sector-field high-resolution ICP-MS. The results showed increasing tungsten levels in all organs with increased dose of exposure, with the highest concentration found in the bones and the lowest concentration found in brain tissue. Gender differences were noticed only in the spleen (higher concentration of tungsten in female animals), and increasing tungsten levels in this organ were correlated with increased iron levels, something that was not observed for any other organ or either of the two other metals analyzed (nickel and cobalt). These findings confirmed most of what has been published on tungsten tissue distribution; they also showed that the brain is relatively protected from oral exposure. Further studies are necessary to clarify the findings in splenic tissue, focusing on possible immunological effects of tungsten exposure.
Assuntos
Compostos de Tungstênio/farmacocinética , Administração Oral , Animais , Cobalto/análise , Cobalto/metabolismo , Cobalto/toxicidade , Feminino , Ferro/análise , Ferro/metabolismo , Ferro/toxicidade , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos C57BL , Níquel/análise , Níquel/metabolismo , Níquel/toxicidade , Distribuição Tecidual , Compostos de Tungstênio/administração & dosagem , Compostos de Tungstênio/toxicidadeRESUMO
Increasing evidence that microRNAs (miRNAs) play important roles in the immune response against infectious agents suggests that miRNA might be exploitable as signatures of exposure to specific infectious agents. In order to identify potential early miRNA biomarkers of bacterial infections, human peripheral blood mononuclear cells (hPBMCs) were exposed to two select agents, Burkholderia pseudomallei K96243 and Francisella tularensis SHU S4, as well as to the nonpathogenic control Escherichia coli DH5α. RNA samples were harvested at three early time points, 30, 60, and 120 minutes postexposure, then sequenced. RNAseq analyses identified 87 miRNAs to be differentially expressed (DE) in a linear fashion. Of these, 31 miRNAs were tested using the miScript miRNA qPCR assay. Through RNAseq identification and qPCR validation, we identified differentially expressed miRNA species that may be involved in the early response to bacterial infections. Based upon its upregulation at early time points postexposure in two different individuals, hsa-mir-30c-5p is a miRNA species that could be studied further as a potential biomarker for exposure to these gram-negative intracellular pathogens. Gene ontology functional analyses demonstrated that programmed cell death is the first ranking biological process associated with miRNAs that are upregulated in F. tularensis-exposed hPBMCs.
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Mosquitoes are efficient, militarily relevant vectors of infectious disease pathogens, including many RNA viruses. The vast majority of all viruses are thought to be undiscovered. Accordingly, recent studies have shown that viruses discovered in insects are very divergent from known pathogens and that many of them lack appropriate reference sequences in the public databases. Given that the majority of viruses are likely still undiscovered, environ mental sampling stands to provide much needed reference samples as well as genetic sequences for comparison. In this study, we sought to determine whether samples of mosquitoes collected from different sites (the Caribbean and locations on the US East Coast) could be differentiated using metagenomic analysis of the RNA viral fraction. We report here distinct virome profiles, even from samples collected short distances apart. In addition to profiling the previously known viruses from these samples, we detected a number of viruses that have been previously undiscovered.
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BACKGROUND: Historically, identification of causal agents of disease has relied heavily on the ability to culture the organism in the laboratory and/or the use of pathogen-specific antibodies or sequence-based probes. However, these methods can be limiting: Even highly sensitive PCR-based assays must be continually updated due to signature degradation as new target strains and near neighbors are sequenced. Thus, there has been a need for assays that do not suffer as greatly from these limitations and/or biases. Recent advances in library preparation technologies for Next-Generation Sequencing (NGS) are focusing on the use of targeted amplification and targeted enrichment/capture to ensure that the most highly discriminating regions of the genomes of known targets (organism-unique regions and/or regions containing functionally important genes or phylogenetically-discriminating SNPs) will be sequenced, regardless of the complex sample background. RESULTS: In the present study, we have assessed the feasibility of targeted sequence enhancement via amplification to facilitate detection of a bacterial pathogen present in low copy numbers in a background of human genomic material. Our results indicate that the targeted amplification of signature regions can effectively identify pathogen genomic material present in as little as 10 copies per ml in a complex sample. Importantly, the correct species and strain calls could be made in amplified samples, while this was not possible in unamplified samples. CONCLUSIONS: The results presented here demonstrate the efficacy of a targeted amplification approach to biothreat detection, using multiple highly-discriminative amplicons per biothreat organism that provide redundancy in case of variation in some primer regions. Importantly, strain level discrimination was possible at levels of 10 genome equivalents. Similar results could be obtained through use of panels focused on the identification of amplicons targeted for specific genes or SNPs instead of, or in addition to, those targeted for specific organisms (ongoing gene-targeting work to be reported later). Note that without some form of targeted enhancement, the enormous background present in complex clinical and environmental samples makes it highly unlikely that sufficient coverage of key pathogen(s) present in the sample will be achieved with current NGS technology to guarantee that the most highly discriminating regions will be sequenced.
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Biblioteca Gênica , Genoma Bacteriano/genética , Genoma Humano/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Análise de Sequência de DNA/métodos , HumanosRESUMO
Historically, cholera outbreaks have been linked to V. cholerae O1 serogroup strains or its derivatives of the O37 and O139 serogroups. A genomic study on the 2010 Haiti cholera outbreak strains highlighted the putative role of non O1/non-O139 V. cholerae in causing cholera and the lack of genomic sequences of such strains from around the world. Here we address these gaps by scanning a global collection of V. cholerae strains as a first step towards understanding the population genetic diversity and epidemic potential of non O1/non-O139 strains. Whole Genome Mapping (Optical Mapping) based bar coding produces a high resolution, ordered restriction map, depicting a complete view of the unique chromosomal architecture of an organism. To assess the genomic diversity of non-O1/non-O139 V. cholerae, we applied a Whole Genome Mapping strategy on a well-defined and geographically and temporally diverse strain collection, the Sakazaki serogroup type strains. Whole Genome Map data on 91 of the 206 serogroup type strains support the hypothesis that V. cholerae has an unprecedented genetic and genomic structural diversity. Interestingly, we discovered chromosomal fusions in two unusual strains that possess a single chromosome instead of the two chromosomes usually found in V. cholerae. We also found pervasive chromosomal rearrangements such as duplications and indels in many strains. The majority of Vibrio genome sequences currently in public databases are unfinished draft sequences. The Whole Genome Mapping approach presented here enables rapid screening of large strain collections to capture genomic complexities that would not have been otherwise revealed by unfinished draft genome sequencing and thus aids in assembling and finishing draft sequences of complex genomes. Furthermore, Whole Genome Mapping allows for prediction of novel V. cholerae non-O1/non-O139 strains that may have the potential to cause future cholera outbreaks.
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Mapeamento Cromossômico/métodos , Variação Genética , Genoma Bacteriano , Vibrio cholerae/genética , Cromossomos Bacterianos/genética , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Duplicação Gênica , Rearranjo Gênico/genética , Tamanho do Genoma , Mutação INDEL/genética , Filogenia , Reprodutibilidade dos Testes , Mapeamento por Restrição , Análise de Sequência de DNARESUMO
BACKGROUND: Understanding the biological roles of microRNAs (miRNAs) is a an active area of research that has produced a surge of publications in PubMed, particularly in cancer research. Along with this increasing interest, many open-source bioinformatics tools to identify existing and/or discover novel miRNAs in next-generation sequencing (NGS) reads become available. While miRNA identification and discovery tools are significantly improved, the development of miRNA differential expression analysis tools, especially in temporal studies, remains substantially challenging. Further, the installation of currently available software is non-trivial and steps of testing with example datasets, trying with one's own dataset, and interpreting the results require notable expertise and time. Subsequently, there is a strong need for a tool that allows scientists to normalize raw data, perform statistical analyses, and provide intuitive results without having to invest significant efforts. FINDINGS: We have developed miRNA Temporal Analyzer (mirnaTA), a bioinformatics package to identify differentially expressed miRNAs in temporal studies. mirnaTA is written in Perl and R (Version 2.13.0 or later) and can be run across multiple platforms, such as Linux, Mac and Windows. In the current version, mirnaTA requires users to provide a simple, tab-delimited, matrix file containing miRNA name and count data from a minimum of two to a maximum of 20 time points and three replicates. To recalibrate data and remove technical variability, raw data is normalized using Normal Quantile Transformation (NQT), and linear regression model is used to locate any miRNAs which are differentially expressed in a linear pattern. Subsequently, remaining miRNAs which do not fit a linear model are further analyzed in two different non-linear methods 1) cumulative distribution function (CDF) or 2) analysis of variances (ANOVA). After both linear and non-linear analyses are completed, statistically significant miRNAs (P < 0.05) are plotted as heat maps using hierarchical cluster analysis and Euclidean distance matrix computation methods. CONCLUSIONS: mirnaTA is an open-source, bioinformatics tool to aid scientists in identifying differentially expressed miRNAs which could be further mined for biological significance. It is expected to provide researchers with a means of interpreting raw data to statistical summaries in a fast and intuitive manner.
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Tungstate (WO²â»4) has been identified as a ground water contaminant at military firing ranges and can be absorbed by ingestion. In this study, C57BL6 mice were exposed to sodium tungstate (Na2WO4·2H2O) (0, 2, 62.5, 125, and 200 mg/kg/day) in their drinking water for an initial 28-day screen and in a one-generation (one-gen) model. Twenty-four hours prior to euthanasia, mice were intraperitoneally injected with Staphylococcal enterotoxin B (SEB) (20 µg/mouse) or saline as controls. After euthanasia, splenocytes and blood were collected and stained with lymphocyte and/or myeloid immunophenotyping panels and analyzed by flow cytometry. In the 28-day and one-gen exposure, statistically significant reductions were observed in the quantities of activated cytotoxic T-cells (TCTL; CD3(+)CD8(+)CD71(+)) and helper T-cells (TH; CD3(+)CD4(+)CD71(+)) from spleens of SEB-treated mice. In the 28-day exposures, CD71(+) TCTL cells were 12.87 ± 2.05% (SE) in the 0 tungstate (control) group compared to 4.44 ± 1.42% in the 200 mg/kg/day (p < 0.001) group. TH cells were 4.85 ± 1.23% in controls and 2.76 ± 0.51% in the 200 mg/kg/day (p < 0.003) group. In the one-gen exposures, TCTL cells were 7.98 ± 0.49% and 6.33 ± 0.49% for P and F1 mice after 0 mg/kg/day tungstate vs 1.58 ± 0.23% and 2.52 ± 0.25% after 200 mg/kg/day of tungstate (p < 0.001). Similarly, TH cells were reduced to 6.21 ± 0.39% and 7.20 ± 0.76%, respectively, for the 0 mg/kg/day P and F1 mice, and 2.28 ± 0.41% and 2.85 ± 0.53%, respectively, for the 200 mg/kg/day tungstate P and F1 groups (p < 0.001). In delayed-type hypersensitivity Type IV experiments, tungstate exposure prior to primary and secondary antigen challenge significantly reduced footpad swelling at 20 and 200 mg/kg/day. These data indicate that exposure to tungstate can result in immune suppression that may, in turn, reduce host defense against pathogens.
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Imunidade Adaptativa/efeitos dos fármacos , Compostos de Tungstênio/farmacologia , Administração Oral , Animais , Enterotoxinas , Feminino , Hipersensibilidade Tardia/imunologia , Imunofenotipagem , Interferon gama/biossíntese , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Linfócitos T Citotóxicos/imunologiaRESUMO
We present draft genome sequences of 15 clinical Vibrio isolates of various serogroups. These are valuable data for use in studying Vibrio cholerae genetic diversity, epidemic potential, and strain attribution.
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BACKGROUND: The development of Next Generation Sequencing (NGS) during the last decade has created an unprecedented amount of sequencing data, as well as the ability to rapidly sequence specimens of interest. Read-based BLAST analysis of NGS data is a common procedure especially in the case of metagenomic samples. However, coverage is usually not enough to allow for de novo assembly. This type of read-based analysis often creates the question of how the reads that align to the same sequence are distributed. The same question applies to preparation of primers or probes for microarray experiments. Although there are several packages that allow the visualization of DNA segments in relation to a reference, in most cases they require the visualization of one reference at a time and the capture of screen shots for each segment. Such a procedure could be tedious and time consuming. The field is in need of a solution that automates the capture of coverage plots for all the segments of interest. RESULTS: We have created BLASTPLOT, a PERL module to quickly plot the BLAST results from short sequences (primers, probes, reads) against reference targets. CONCLUSIONS: BLASTPLOT is a simple to use PERL module that allows the generation of PNG graphs for all the reference sequences associated with a BLAST result set.
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Middle East respiratory syndrome coronavirus (MERS-CoV) is the etiologic agent of a highly lethal pneumonia. Here, we report the full-genome sequence of the Jordan-N3/2012 strain after serial passage in two distinct mammalian cell lines. The genome exhibits noteworthy stability, which may inform the development of vaccines and therapeutics used to treat infection with this virus.
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BACKGROUND: The availability of genetic data has increased dramatically in recent years. The greatest value of this data is its potential for personalized medicine. Many new associations are reported every day from Genome Wide Association Studies (GWAS). However, robust, reproducible associations are elusive for some complex diseases. Ontologies present a potential way to distinguish between spurious associations and those with a potential influence on the phenotype. Such an approach would be based on finding associations of the same genetic variant with closely related, but distinct, phenotypes. This approach can be accomplished with a phenotype ontology that also holds genetic association data. RESULTS: Here, we report a structured knowledge application to navigate and to facilitate the discovery of relationships between different phenotypes and their genetic associations. CONCLUSIONS: OGA allows users to (1) find the intersecting set of genes for phenotypes of interest, (2) find empirical p values for such observations and (3) OGA outperforms similar applications in number of total concepts and genes mapped.
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Estudo de Associação Genômica Ampla , Genótipo , Fenótipo , Software , Predisposição Genética para Doença , Genoma Humano , Humanos , Anotação de Sequência MolecularRESUMO
Rickettsia prowazekii is a notable intracellular pathogen, the agent of epidemic typhus, and a potential biothreat agent. We present here whole-genome sequence data for four strains of R. prowazekii, including one from a flying squirrel.