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pathogen dna enrichment

Xiao-Yu Hu, Yu-Fei Yao, Bo-Miao Cui, Jun Lv, Xin Shen, Biao Ren, Ming-Yun Li, Qiang Guo, Rui-Jie Huang, Yan Li
OBJECTIVE: To analyze the whole microbial structure in a case of rampant caries to provide evidence for its prevention and treatment. METHODS: Clinical samples including blood, supragingival plaque, plaque in the caries cavity, saliva, and mucosal swabs were collected with the patient's consent. The blood sample was sent for routine immune test, and the others samples were stained using Gram method and cultured for identifying colonies and 16S rRNA sequencing. DNA was extracted from the samples and tested for the main cariogenic bacterium (Streptococcus mutans) with qPCR, and the whole microbial structure was analyzed using DGGE...
October 20, 2016: Nan Fang Yi Ke da Xue Xue Bao, Journal of Southern Medical University
Eva Gross, Harm van Tinteren, Zhou Li, Sandra Raab, Christina Meul, Stefanie Avril, Nadja Laddach, Michaela Aubele, Corinna Propping, Apostolos Gkazepis, Manfred Schmitt, Alfons Meindl, Petra M Nederlof, Marion Kiechle, Esther H Lips
BACKGROUND: Triple-negative breast cancer (TNBC) with a BRCA1-like molecular signature has been demonstrated to remarkably respond to platinum-based chemotherapy and might be suited for a future treatment with poly(ADP-ribose)polymerase (PARP) inhibitors. In order to rapidly assess this signature we have previously developed a multiplex-ligation-dependent probe amplification (MLPA)-based assay. Here we present an independent validation of this assay to confirm its important clinical impact...
October 19, 2016: BMC Cancer
Juan David Ospina-Villa, Absalom Zamorano-Carrillo, Carlos A Castañon-Sanchez, Esther Ramirez-Moreno, Laurence A Marchat
Aptamers are short single-stranded RNA or DNA oligonucleotides that are capable of binding various biological targets with high affinity and specificity. Their identification initially relies on a molecular process named SELEX (Systematic Evolution of Ligands by EXponential enrichment) that has been later modified in order to improve aptamer sensitivity, minimize duration and cost of the assay, as well as increase target types. Several biochemical modifications can help to enhance aptamer stability without affecting significantly target interaction...
October 15, 2016: Brazilian Journal of Infectious Diseases
Utsav Pandey, Andrew S Bell, Daniel W Renner, David A Kennedy, Jacob T Shreve, Chris L Cairns, Matthew J Jones, Patricia A Dunn, Andrew F Read, Moriah L Szpara
The intensification of the poultry industry over the last 60 years facilitated the evolution of increased virulence and vaccine breaks in Marek's disease virus (MDV-1). Full-genome sequences are essential for understanding why and how this evolution occurred, but what is known about genome-wide variation in MDV comes from laboratory culture. To rectify this, we developed methods for obtaining high-quality genome sequences directly from field samples without the need for sequence-based enrichment strategies prior to sequencing...
September 2016: MSphere
Lucia Rochi, María José Diéguez, Germán Burguener, Martín Alejandro Darino, María Fernanda Pergolesi, Lorena Romina Ingala, Alba Romina Cuyeu, Adrián Turjanski, Enrique Domingo Kreff, Francisco Sacco
Rust fungi are one of the most devastating pathogens of crop plants. The biotrophic fungus Puccinia sorghi Schwein (Ps) is responsible for maize common rust, an endemic disease of maize (Zea mays L) in Argentina that causes significant yield losses in corn production. In spite of this, the Ps genomic sequence was not available. We used Illumina sequencing to rapidly produce the 99.6 Mb draft genome sequence of Ps race RO10H11247, derived from a single-uredinial isolate from infected maize leaves collected in the Argentine Corn Belt Region during 2010...
October 13, 2016: Fungal Genetics and Biology: FG & B
Helen D Donoghue
Tuberculosis is a significant global disease today, so understanding its origins and history is important. It is primarily a lung infection and is transmitted by infectious aerosols from person to person, so a high population density encourages its spread. The causative organism is Mycobacterium tuberculosis, an obligate pathogen in the M. tuberculosis complex that also contains closely related species, such as Mycobacterium bovis, that primarily infect animals. Typical bone lesions occur in about 5% of untreated infections...
August 2016: Microbiology Spectrum
J Stephen Dumler, Sara H Sinclair, Valeria Pappas-Brown, Amol C Shetty
Anaplasma phagocytophilum, an obligate intracellular prokaryote, infects neutrophils, and alters cardinal functions via reprogrammed transcription. Large contiguous regions of neutrophil chromosomes are differentially expressed during infection. Secreted A. phagocytophilum effector AnkA transits into the neutrophil or granulocyte nucleus to complex with DNA in heterochromatin across all chromosomes. AnkA binds to gene promoters to dampen cis-transcription and also has features of matrix attachment region (MAR)-binding proteins that regulate three-dimensional chromatin architecture and coordinate transcriptional programs encoded in topologically-associated chromatin domains...
2016: Frontiers in Cellular and Infection Microbiology
Rowan R Thomas, Heather J L Brooks, Rory O'Brien
BACKGROUND: Shiga toxin-producing (STEC) and enteropathogenic (EPEC) Escherichia coli are gastrointestinal pathogens causing diarrhoeal and extraintestinal disease. Due to lack of EPEC screening and use of Sorbitol-MacConkey (SMAC) agar in faecal screening, the true prevalence of EPEC and non-O157 STEC in New Zealand diarrhoeal cases is unknown. METHODS: Diarrhoeic stools sourced from Dunedin hospital were pre-enriched, DNA extracted with Chelex-100 resin and screened using a multiplex TaqMan quantitative PCR assay amplifying stx1, sxt2 and EPEC (eae) gene markers...
October 3, 2016: Journal of Clinical Pathology
Jun Teng, Fang Yuan, Yingwang Ye, Lei Zheng, Li Yao, Feng Xue, Wei Chen, Baoguang Li
Aptamers are single stranded DNA or RNA ligands, which can be selected by a method called systematic evolution of ligands by exponential enrichment (SELEX); and they can specifically recognize and bind to their targets. These unique characteristics of aptamers offer great potentials in applications such as pathogen detection and biomolecular screening. Pathogen detection is the critical means in detecting and identifying the problems related to public health and food safety; and only the rapid, sensitive and efficient detection technologies can enable the users to make the accurate assessments on the risks of infections (humans and animals) or contaminations (foods and other commodities) caused by various pathogens...
2016: Frontiers in Microbiology
Charlotte J Houldcroft, Josephine M Bryant, Daniel P Depledge, Ben K Margetts, Jacob Simmonds, Stephanos Nicolaou, Helena J Tutill, Rachel Williams, Austen J J Worth, Stephen D Marks, Paul Veys, Elizabeth Whittaker, Judith Breuer
Human cytomegalovirus (HCMV) is a significant pathogen in immunocompromised individuals, with the potential to cause fatal pneumonitis and colitis, as well as increasing the risk of organ rejection in transplant patients. With the advent of new anti-HCMV drugs there is therefore considerable interest in using virus sequence data to monitor emerging resistance to antiviral drugs in HCMV viraemia and disease, including the identification of putative new mutations. We used target-enrichment to deep sequence HCMV DNA from 11 immunosuppressed pediatric patients receiving single or combination anti-HCMV treatment, serially sampled over 1-27 weeks...
2016: Frontiers in Microbiology
K Schmidt, S Mwaigwisya, L C Crossman, M Doumith, D Munroe, C Pires, A M Khan, N Woodford, N J Saunders, J Wain, J O'Grady, D M Livermore
OBJECTIVES: The introduction of metagenomic sequencing to diagnostic microbiology has been hampered by slowness, cost and complexity. We explored whether MinION nanopore sequencing could accelerate diagnosis and resistance profiling, using complicated urinary tract infections as an exemplar. METHODS: Bacterial DNA was enriched from clinical urines (n = 10) and from healthy urines 'spiked' with multiresistant Escherichia coli (n = 5), then sequenced by MinION...
September 25, 2016: Journal of Antimicrobial Chemotherapy
Lucas Nojosa Oliveira, Luciana Casaletti, Sônia Nair Báo, Clayton Luiz Borges, Patrícia de Sousa Lima, Célia Maria de Almeida Soares
Paracoccidioidomycosis is an endemic disease in Latin America, caused by thermo dimorphic fungi of the genus Paracoccidioides. Although previous proteome analyses of Paracoccidioides spp. have been carried out, the nuclear subproteome of this pathogen has not been described. In this way, we aimed to characterize the nuclear proteome of Paracoccidioides species, in the yeast form. For that, yeast cells were disrupted and submitted to cell fractionation. The purity of the nuclear fraction was confirmed by fluorescence and electron microscopy...
October 2016: Fungal Biology
Helen Auburn, Mark Zuckerman, Melvyn Smith
AbstractIn order to develop novel host/pathogen real- time PCR assays for routine diagnostic use, early gene expression patterns from both Epstein-Barr virus (EBV) and Raji cells were examined after inducing the lytic life cycle using 12-O-tetradecanoyl-13-phorbol ester (TPA) and sodium butyrate. Real-time PCR identified several highly induced (>90-fold) EBV lytic genes over a 48 hr time-course during the lytic induction phase. Latent genes were induced at low levels during this phase. The cellular response to lytic viral replication is poorly understood...
September 12, 2016: Journal of Medical Microbiology
Alberto Ortiz, Holger Husi, Laura Gonzalez-Lafuente, Lara Valiño-Rivas, Manuel Fresno, Ana Belen Sanz, William Mullen, Amaya Albalat, Sergio Mezzano, Tonia Vlahou, Harald Mischak, Maria Dolores Sanchez-Niño
An improved understanding of pathogenic pathways in AKI may identify novel therapeutic approaches. Previously, we conducted unbiased liquid chromatography-tandem mass spectrometry-based protein expression profiling of the renal proteome in mice with acute folate nephropathy. Here, analysis of the dataset identified enrichment of pathways involving NFκB in the kidney cortex, and a targeted data mining approach identified components of the noncanonical NFκB pathway, including the upstream kinase mitogen-activated protein kinase kinase kinase 14 (MAP3K14), the NFκB DNA binding heterodimer RelB/NFκB2, and proteins involved in NFκB2 p100 ubiquitination and proteasomal processing to p52, as upregulated...
September 12, 2016: Journal of the American Society of Nephrology: JASN
Mélanie Massonnet, Abraham Morales-Cruz, Rosa Figueroa-Balderas, Daniel P Lawrence, Kendra Baumgartner, Dario Cantu
The ascomycete Neofusicoccum parvum, one of the causal agents of Botryosphaeria dieback, is a destructive wood-infecting fungus and a serious threat to grape production worldwide. The capability of colonizing woody tissue combined with the secretion of phytotoxic compounds is thought to underlie its pathogenicity and virulence. Here, we describe the repertoire of virulence factors and their transcriptional dynamics as the fungus feeds on different substrates and colonizes the woody stem. We assembled and annotated a highly contiguous genome using single molecule real-time DNA sequencing...
September 8, 2016: Molecular Plant Pathology
Giulia Amagliani, Annalisa Petruzzelli, Elisa Carloni, Franco Tonucci, Martina Foglini, Eleonora Micci, Mariagrazia Ricci, Stefania Di Lullo, Luca Rotundo, Giorgio Brandi
Italy is one of the main producers and exporters of cheese made from unpasteurized sheep milk. Since raw milk and its derived products are known sources of human infections, cheese produced from raw sheep milk could pose a microbiological threat to public health. Hence, the objectives of the study were: to characterize the potential risk of the presence of pathogens Escherichia coli O157, Listeria monocytogenes, and Salmonella in raw ovine milk destined for cheese production obtained from all the sheep farms (n = 24) in the Marches region (Central Italy) that directly transform raw milk into cheeses and to evaluate the equivalence between the analytical methods applied...
September 8, 2016: Foodborne Pathogens and Disease
Jacob L Steenwyk, John S Soghigian, John R Perfect, John G Gibbons
BACKGROUND: Copy number variants (CNVs) are a class of structural variants (SVs) and are defined as fragments of DNA that are present at variable copy number in comparison with a reference genome. Recent advances in bioinformatics methodologies and sequencing technologies have enabled the high-resolution quantification of genome-wide CNVs. In pathogenic fungi SVs have been shown to alter gene expression, influence host specificity, and drive fungicide resistance, but little attention has focused specifically on CNVs...
2016: BMC Genomics
Douglas R Deutsch, Bryan Utter, Vincent A Fischetti
Staphylococcus aureus is a major clinically important pathogen with well-studied phage contributions to its virulence potential. In this commentary, we describe our method to enrich and sequence stealth extra-chromosomal DNA elements in the bacterial cell, allowing the identification of novel extra-chromosomal prophages in S. aureus clinical strains. Extra-chromosomal sequencing is a useful and broadly applicable tool to study bacterial genomics, giving a temporal glance at the extra-chromosomal compartment of the cell and allowing researchers to uncover lower-copy plasmidial elements (e...
July 2016: Mobile Genetic Elements
Claudio Donati, Moreno Zolfo, Davide Albanese, Duy Tin Truong, Francesco Asnicar, Valerio Iebba, Duccio Cavalieri, Olivier Jousson, Carlotta De Filippo, Curtis Huttenhower, Nicola Segata
Microbial epidemiology and population genomics have previously been carried out near-exclusively for organisms grown in vitro. Metagenomics helps to overcome this limitation, but it is still challenging to achieve strain-level characterization of microorganisms from culture-independent data with sufficient resolution for epidemiological modelling. Here, we have developed multiple complementary approaches that can be combined to profile and track individual microbial strains. To specifically profile highly recombinant neisseriae from oral metagenomes, we integrated four metagenomic analysis techniques: single nucleotide polymorphisms in the clade's core genome, DNA uptake sequence signatures, metagenomic multilocus sequence typing and strain-specific marker genes...
2016: Nature Microbiology
Jarrett D Morrow, Michael H Cho, Craig P Hersh, Victor Pinto-Plata, Bartolome Celli, Nathaniel Marchetti, Gerard Criner, Raphael Bueno, George Washko, Kimberly Glass, Augustine M K Choi, John Quackenbush, Edwin K Silverman, Dawn L DeMeo
Chronic obstructive pulmonary disease (COPD) is a smoking-related disease characterized by genetic and phenotypic heterogeneity. Although association studies have identified multiple genomic regions with replicated associations to COPD, genetic variation only partially explains the susceptibility to lung disease, and suggests the relevance of epigenetic investigations. We performed genome-wide DNA methylation profiling in homogenized lung tissue samples from 46 control subjects with normal lung function and 114 subjects with COPD, all former smokers...
August 26, 2016: Epigenetics: Official Journal of the DNA Methylation Society
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