Chloroform-Treated Filamentous Phage as a Bioreceptor for Piezoelectric Sensors

National Research Council Canada - National Science Library

Olsen, Eric V; Sykora, Jennifer C; Sorokulova, Iryna B; Petrenko, Valery A; Chen, I-Hsuan; Barbaree, James M; Vodyanoy, Vitaly J

2005-01-01

... and S. typhimurium. ELISA confirmed affinity-selected phage specificity for streptavidin. Spheroid induction was optimized to achieve greatest conversion yields as a function of solvent exposure time and concentration...

Targeting Antibacterial Agents by Using Drug-Carrying Filamentous Bacteriophages

Science.gov (United States)

Yacoby, Iftach; Shamis, Marina; Bar, Hagit; Shabat, Doron; Benhar, Itai

2006-01-01

Bacteriophages have been used for more than a century for (unconventional) therapy of bacterial infections, for half a century as tools in genetic research, for 2 decades as tools for discovery of specific target-binding proteins, and for nearly a decade as tools for vaccination or as gene delivery vehicles. Here we present a novel application of filamentous bacteriophages (phages) as targeted drug carriers for the eradication of (pathogenic) bacteria. The phages are genetically modified to display a targeting moiety on their surface and are used to deliver a large payload of a cytotoxic drug to the target bacteria. The drug is linked to the phages by means of chemical conjugation through a labile linker subject to controlled release. In the conjugated state, the drug is in fact a prodrug devoid of cytotoxic activity and is activated following its dissociation from the phage at the target site in a temporally and spatially controlled manner. Our model target was Staphylococcus aureus, and the model drug was the antibiotic chloramphenicol. We demonstrated the potential of using filamentous phages as universal drug carriers for targetable cells involved in disease. Our approach replaces the selectivity of the drug itself with target selectivity borne by the targeting moiety, which may allow the reintroduction of nonspecific drugs that have thus far been excluded from antibacterial use (because of toxicity or low selectivity). Reintroduction of such drugs into the arsenal of useful tools may help to combat emerging bacterial antibiotic resistance. PMID:16723570

Phage-display libraries of murine and human antibody Fab fragments

DEFF Research Database (Denmark)

Engberg, J; Andersen, P S; Nielsen, L K

1996-01-01

We provide efficient and detailed procedures for construction, expression, and screening of comprehensive libraries of murine or human antibody Fab fragments displayed on the surface of filamentous phage. In addition, protocols for producing and using ultra-electrocompetent cells, for producing Fab...

Inhibition of bacterial conjugation by phage M13 and its protein g3p: quantitative analysis and model.

Directory of Open Access Journals (Sweden)

Abraham Lin

Full Text Available Conjugation is the main mode of horizontal gene transfer that spreads antibiotic resistance among bacteria. Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances. Filamentous bacteriophages were first observed to inhibit conjugation several decades ago. Here we investigate the mechanism of inhibition and find that the primary effect on conjugation is occlusion of the conjugative pilus by phage particles. This interaction is mediated primarily by phage coat protein g3p, and exogenous addition of the soluble fragment of g3p inhibited conjugation at low nanomolar concentrations. Our data are quantitatively consistent with a simple model in which association between the pili and phage particles or g3p prevents transmission of an F plasmid encoding tetracycline resistance. We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration. Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage, these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

Membrane insertion and assembly of epitope-tagged gp9 at the tip of the M13 phage

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Kuhn Andreas

2011-09-01

Full Text Available Abstract Background Filamentous M13 phage extrude from infected Escherichia coli with a tip structure composed of gp7 and gp9. This tip structure is extended by the assembly of the filament composed of the major coat protein gp8. Finally, gp3 and gp6 terminate the phage structure at the proximal end. Up to now, gp3 has been the primary tool for phage display technology. However, gp7, gp8 and gp9 could also be used for phage display and these phage particles should bind to two different or more surfaces when the modified coat proteins are combined. Therefore, we tested here if the amino-terminal end of gp9 can be modified and whether the modified portion is exposed and detectable on the M13 phage particles. Results The amino-terminal region of gp9 was modified by inserting short sequences that encode antigenic epitopes. We show here that the modified gp9 proteins correctly integrate into the membrane using the membrane insertase YidC exposing the modified epitope into the periplasm. The proteins are then efficiently assembled onto the phage particles. Also extensions up to 36 amino acid residues at the amino-terminal end of gp9 did not interfere with membrane integration and phage assembly. The exposure of the antigenic tags on the phage was visualised with immunogold labelling by electron microscopy and verified by dot blotting with antibodies to the tags. Conclusions Our results suggest that gp9 at the phage tip is suitable for the phage display technology. The modified gp9 can be supplied in trans from a plasmid and fully complements M13 phage with an amber mutation in gene 9. The modified phage tip is very well accessible to antibodies.

Targeting Antibacterial Agents by Using Drug-Carrying Filamentous Bacteriophages

OpenAIRE

Yacoby, Iftach; Shamis, Marina; Bar, Hagit; Shabat, Doron; Benhar, Itai

2006-01-01

Bacteriophages have been used for more than a century for (unconventional) therapy of bacterial infections, for half a century as tools in genetic research, for 2 decades as tools for discovery of specific target-binding proteins, and for nearly a decade as tools for vaccination or as gene delivery vehicles. Here we present a novel application of filamentous bacteriophages (phages) as targeted drug carriers for the eradication of (pathogenic) bacteria. The phages are genetically modified to d...

Genetic engineering of a temperate phage-based delivery system for CRISPR/Cas9 antimicrobials against Staphylococcus aureus.

Science.gov (United States)

Park, Joo Youn; Moon, Bo Youn; Park, Juw Won; Thornton, Justin A; Park, Yong Ho; Seo, Keun Seok

2017-03-21

Discovery of clustered, regularly interspaced, short palindromic repeats and the Cas9 RNA-guided nuclease (CRISPR/Cas9) system provides a new opportunity to create programmable gene-specific antimicrobials that are far less likely to drive resistance than conventional antibiotics. However, the practical therapeutic use of CRISPR/Cas9 is still questionable due to current shortcomings in phage-based delivery systems such as inefficient delivery, narrow host range, and potential transfer of virulence genes by generalized transduction. In this study, we demonstrate genetic engineering strategies to overcome these shortcomings by integrating CRISPR/Cas9 system into a temperate phage genome, removing major virulence genes from the host chromosome, and expanding host specificity of the phage by complementing tail fiber protein. This significantly improved the efficacy and safety of CRISPR/Cas9 antimicrobials to therapeutic levels in both in vitro and in vivo assays. The genetic engineering tools and resources established in this study are expected to provide an efficacious and safe CRISPR/Cas9 antimicrobial, broadly applicable to Staphylococcus aureus.

A virulence-associated filamentous bacteriophage of Neisseria meningitidis increases host-cell colonisation.

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Emmanuelle Bille

2017-07-01

Full Text Available Neisseria meningitidis is a commensal of human nasopharynx. In some circumstances, this bacteria can invade the bloodstream and, after crossing the blood brain barrier, the meninges. A filamentous phage, designated MDAΦ for Meningococcal Disease Associated, has been associated with invasive disease. In this work we show that the prophage is not associated with a higher virulence during the bloodstream phase of the disease. However, looking at the interaction of N. meningitidis with epithelial cells, a step essential for colonization of the nasopharynx, we demonstrate that the presence of the prophage, via the production of viruses, increases colonization of encapsulated meningococci onto monolayers of epithelial cells. The analysis of the biomass covering the epithelial cells revealed that meningococci are bound to the apical surface of host cells by few layers of heavily piliated bacteria, whereas, in the upper layers, bacteria are non-piliated but surrounded by phage particles which (i form bundles of filaments, and/or (ii are in some places associated with bacteria. The latter are likely to correspond to growing bacteriophages during their extrusion through the outer membrane. These data suggest that, as the biomass increases, the loss of piliation in the upper layers of the biomass does not allow type IV pilus bacterial aggregation, but is compensated by a large production of phage particles that promote bacterial aggregation via the formation of bundles of phage filaments linked to the bacterial cell walls. We propose that MDAΦ by increasing bacterial colonization in the mucosa at the site-of-entry, increase the occurrence of diseases.

Affinity-Selected Filamentous Bacteriophage as a Probe for Acoustic Wave Biodetectors of Salmonella typhimurium

National Research Council Canada - National Science Library

Olsen, Eric V; Sorokulova, Iryna B; Petrenko, Valery A; Chen, I-Hsuan; Barbaree, James M; Vodyanoy, Vitaly J

2005-01-01

Proof-in-concept biosensors were prepared for the rapid detection of Salmonella typhimurium in solution, based on affinity-selected filamentous phage prepared as probes physically adsorbed to piezoelectric transducers...

A century of the phage: past, present and future.

Science.gov (United States)

Salmond, George P C; Fineran, Peter C

2015-12-01

Viruses that infect bacteria (bacteriophages; also known as phages) were discovered 100 years ago. Since then, phage research has transformed fundamental and translational biosciences. For example, phages were crucial in establishing the central dogma of molecular biology - information is sequentially passed from DNA to RNA to proteins - and they have been shown to have major roles in ecosystems, and help drive bacterial evolution and virulence. Furthermore, phage research has provided many techniques and reagents that underpin modern biology - from sequencing and genome engineering to the recent discovery and exploitation of CRISPR-Cas phage resistance systems. In this Timeline, we discuss a century of phage research and its impact on basic and applied biology.

Corruption of phage-display libraries by target-unrelated clones: Diagnosis and countermeasures

Science.gov (United States)

Thomas, William D.; Golomb, Miriam; Smith, George P.

2010-01-01

Phage display is used to discover peptides or proteins with a desired target property—most often, affinity for a target selector molecule. Libraries of phage clones displaying diverse surface peptides are subject to a selection process designed to enrich for the target behavior, and subsequently propagated to restore phage numbers. A recurrent problem is enrichment of clones, called target-unrelated phage (TUPs), that lack the target behavior. Many TUPs are propagation-related; they have mutations conferring a growth advantage, and are enriched during the propagations accompanying selection. Unlike other filamentous phage libraries, fd-tet-based libraries are relatively resistant to propagation-related TUP corruption. Their minus strand origin is disrupted by a large cassette that simultaneously confers resistance to tetracycline and imposes a rate-limiting growth defect that cannot be bypassed with simple mutations. Nonetheless, a new type of propagation-related TUP emerged in the output of in vivo selections from an fd-tet library. The founding clone had a complex rearrangement that restored the minus strand origin while retaining tetracycline resistance. The rearrangement involved two recombination events, one with a contaminant having a wild-type minus strand origin. The founder’s infectivity advantage spread by simple recombination to clones displaying different peptides. We propose measures for minimizing TUP corruption. PMID:20692225

Ligand-directed profiling of organelles with internalizing phage libraries

Science.gov (United States)

Dobroff, Andrey S.; Rangel, Roberto; Guzman-Roja, Liliana; Salmeron, Carolina C.; Gelovani, Juri G.; Sidman, Richard L.; Bologa, Cristian G.; Oprea, Tudor I.; Brinker, C. Jeffrey; Pasqualini, Renata; Arap, Wadih

2015-01-01

Phage display is a resourceful tool to, in an unbiased manner, discover and characterize functional protein-protein interactions, to create vaccines, and to engineer peptides, antibodies, and other proteins as targeted diagnostic and/or therapeutic agents. Recently, our group has developed a new class of internalizing phage (iPhage) for ligand-directed targeting of organelles and/or to identify molecular pathways within live cells. This unique technology is suitable for applications ranging from fundamental cell biology to drug development. Here we describe the method for generating and screening the iPhage display system, and explain how to select and validate candidate internalizing homing peptide. PMID:25640897

Viruses Infecting a Freshwater Filamentous Cyanobacterium (Nostoc sp.) Encode a Functional CRISPR Array and a Proteobacterial DNA Polymerase B.

Science.gov (United States)

Chénard, Caroline; Wirth, Jennifer F; Suttle, Curtis A

2016-06-14

Here we present the first genomic characterization of viruses infecting Nostoc, a genus of ecologically important cyanobacteria that are widespread in freshwater. Cyanophages A-1 and N-1 were isolated in the 1970s and infect Nostoc sp. strain PCC 7210 but remained genomically uncharacterized. Their 68,304- and 64,960-bp genomes are strikingly different from those of other sequenced cyanophages. Many putative genes that code for proteins with known functions are similar to those found in filamentous cyanobacteria, showing a long evolutionary history in their host. Cyanophage N-1 encodes a CRISPR array that is transcribed during infection and is similar to the DR5 family of CRISPRs commonly found in cyanobacteria. The presence of a host-related CRISPR array in a cyanophage suggests that the phage can transfer the CRISPR among related cyanobacteria and thereby provide resistance to infection with competing phages. Both viruses also encode a distinct DNA polymerase B that is closely related to those found in plasmids of Cyanothece sp. strain PCC 7424, Nostoc sp. strain PCC 7120, and Anabaena variabilis ATCC 29413. These polymerases form a distinct evolutionary group that is more closely related to DNA polymerases of proteobacteria than to those of other viruses. This suggests that the polymerase was acquired from a proteobacterium by an ancestral virus and transferred to the cyanobacterial plasmid. Many other open reading frames are similar to a prophage-like element in the genome of Nostoc sp. strain PCC 7524. The Nostoc cyanophages reveal a history of gene transfers between filamentous cyanobacteria and their viruses that have helped to forge the evolutionary trajectory of this previously unrecognized group of phages. Filamentous cyanobacteria belonging to the genus Nostoc are widespread and ecologically important in freshwater, yet little is known about the genomic content of their viruses. Here we report the first genomic analysis of cyanophages infecting

Basic Phage Mathematics.

Science.gov (United States)

Abedon, Stephen T; Katsaounis, Tena I

2018-01-01

Basic mathematical descriptions are useful in phage ecology, applied phage ecology such as in the course of phage therapy, and also toward keeping track of expected phage-bacterial interactions as seen during laboratory manipulation of phages. The most basic mathematical descriptor of phages is their titer, that is, their concentration within stocks, experimental vessels, or other environments. Various phenomena can serve to modify phage titers, and indeed phage titers can vary as a function of how they are measured. An important aspect of how changes in titers can occur results from phage interactions with bacteria. These changes tend to vary in degree as a function of bacterial densities within environments, and particularly densities of those bacteria that are susceptible to or at least adsorbable by a given phage type. Using simple mathematical models one can describe phage-bacterial interactions that give rise particularly to phage adsorption events. With elaboration one can consider changes in both phage and bacterial densities as a function of both time and these interactions. In addition, phages along with their impact on bacteria can be considered as spatially constrained processes. In this chapter we consider the simpler of these concepts, providing in particular detailed verbal explanations toward facile mathematical insight. The primary goal is to stimulate a more informed use and manipulation of phages and phage populations within the laboratory as well as toward more effective phage application outside of the laboratory, such as during phage therapy. More generally, numerous issues and approaches to the quantification of phages are considered along with the quantification of individual, ecological, and applied properties of phages.

Corruption of phage display libraries by target-unrelated clones: diagnosis and countermeasures.

Science.gov (United States)

Thomas, William D; Golomb, Miriam; Smith, George P

2010-12-15

Phage display is used to discover peptides or proteins with a desired target property-most often, affinity for a target selector molecule. Libraries of phage clones displaying diverse surface peptides are subject to a selection process designed to enrich for the target behavior and subsequently propagated to restore phage numbers. A recurrent problem is enrichment of clones, called target-unrelated phages or peptides (TUPs), that lack the target behavior. Many TUPs are propagation related; they have mutations conferring a growth advantage and are enriched during the propagations accompanying selection. Unlike other filamentous phage libraries, fd-tet-based libraries are relatively resistant to propagation-related TUP corruption. Their minus-strand origin is disrupted by a large cassette that simultaneously confers resistance to tetracycline and imposes a rate-limiting growth defect that cannot be bypassed with simple mutations. Nonetheless, a new type of propagation-related TUP emerged in the output of in vivo selections from an fd-tet library. The founding clone had a complex rearrangement that restored the minus-strand origin while retaining tetracycline resistance. The rearrangement involved two recombination events, one with a contaminant having a wild-type minus-strand origin. The founder's infectivity advantage spread by simple recombination to clones displaying different peptides. We propose measures for minimizing TUP corruption. Copyright © 2010 Elsevier Inc. All rights reserved.

Synthetic Phage for Tissue Regeneration

Directory of Open Access Journals (Sweden)

So Young Yoo

2014-01-01

Full Text Available Controlling structural organization and signaling motif display is of great importance to design the functional tissue regenerating materials. Synthetic phage, genetically engineered M13 bacteriophage has been recently introduced as novel tissue regeneration materials to display a high density of cell-signaling peptides on their major coat proteins for tissue regeneration purposes. Structural advantages of their long-rod shape and monodispersity can be taken together to construct nanofibrous scaffolds which support cell proliferation and differentiation as well as direct orientation of their growth in two or three dimensions. This review demonstrated how functional synthetic phage is designed and subsequently utilized for tissue regeneration that offers potential cell therapy.

Ff-nano, Short Functionalized Nanorods Derived from Ff (f1, fd or M13 Filamentous Bacteriophage

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Sadia eSattar

2015-04-01

Full Text Available F-specific filamentous phage of Escherichia coli (Ff: f1, M13 or fd are long thin filaments (860 nm x 6 nm. They have been a major workhorse in display technologies and bionanotechnology; however, some applications are limited by the high length-to-diameter ratio of Ff. Furthermore, use of functionalized Ff outside of laboratory containment is in part hampered by the fact that they are genetically modified viruses. We have now developed a system for production and purification of very short functionalized Ff-phage-derived nanorods, named Ff-nano, that are only 50 nm in length. In contrast to standard Ff-derived vectors that replicate in E. coli and contain antibiotic-resistance genes, Ff-nano are protein DNA complexes that cannot replicate on their own and do not contain any coding sequences. These nanorods show an increased resistance to heating at 70 °C in 1 % SDS in comparison to the full-length Ff phage of the same coat composition. We demonstrate that functionalized Ff-nano particles are suitable for application as detection particles in sensitive and quantitative dipstick lateral flow diagnostic assay for human plasma fibronectin.

Hybrid Nanomaterial Complexes for Advanced Phage-guided Gene Delivery

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Teerapong Yata

2014-01-01

Full Text Available Developing nanomaterials that are effective, safe, and selective for gene transfer applications is challenging. Bacteriophages (phage, viruses that infect bacteria only, have shown promise for targeted gene transfer applications. Unfortunately, limited progress has been achieved in improving their potential to overcome mammalian cellular barriers. We hypothesized that chemical modification of the bacteriophage capsid could be applied to improve targeted gene delivery by phage vectors into mammalian cells. Here, we introduce a novel hybrid system consisting of two classes of nanomaterial systems, cationic polymers and M13 bacteriophage virus particles genetically engineered to display a tumor-targeting ligand and carry a transgene cassette. We demonstrate that the phage complex with cationic polymers generates positively charged phage and large aggregates that show enhanced cell surface attachment, buffering capacity, and improved transgene expression while retaining cell type specificity. Moreover, phage/polymer complexes carrying a therapeutic gene achieve greater cancer cell killing than phage alone. This new class of hybrid nanomaterial platform can advance targeted gene delivery applications by bacteriophage.

Expert Opinion on Three Phage Therapy Related Topics: Bacterial Phage Resistance, Phage Training and Prophages in Bacterial Production Strains

Directory of Open Access Journals (Sweden)

Christine Rohde

2018-04-01

Full Text Available Phage therapy is increasingly put forward as a “new” potential tool in the fight against antibiotic resistant infections. During the “Centennial Celebration of Bacteriophage Research” conference in Tbilisi, Georgia on 26–29 June 2017, an international group of phage researchers committed to elaborate an expert opinion on three contentious phage therapy related issues that are hampering clinical progress in the field of phage therapy. This paper explores and discusses bacterial phage resistance, phage training and the presence of prophages in bacterial production strains while reviewing relevant research findings and experiences. Our purpose is to inform phage therapy stakeholders such as policy makers, officials of the competent authorities for medicines, phage researchers and phage producers, and members of the pharmaceutical industry. This brief also points out potential avenues for future phage therapy research and development as it specifically addresses those overarching questions that currently call for attention whenever phages go into purification processes for application.

Synergy as a rationale for phage therapy using phage cocktails.

Science.gov (United States)

Schmerer, Matthew; Molineux, Ian J; Bull, James J

2014-01-01

Where phages are used to treat bacterial contaminations and infections, multiple phages are typically applied at once as a cocktail. When two or more phages in the cocktail attack the same bacterium, the combination may produce better killing than any single phage (synergy) or the combination may be worse than the best single phage (interference). Synergy is of obvious utility, especially if it can be predicted a priori, but it remains poorly documented with few examples known. This study addresses synergy in which one phage improves adsorption by a second phage. It first presents evidence of synergy from an experimental system of two phages and a mucoid E. coli host. The synergy likely stems from a tailspike enzyme produced by one of the phages. We then offer mathematical models and simulations to understand the dynamics of synergy and the enhanced magnitude of bacterial control possible. The models and observations complement each other and suggest that synergy may be of widespread utility and may be predictable from easily observed phenotypes.

Characterization of polyacrylamide-stabilized Pf1 phage liquid crystals for protein NMR spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Trempe, Jean-Francois; Morin, Frederick G.; Xia Zhicheng; Marchessault, Robert H.; Gehring, Kalle [McGill University, Department of Biochemistry and Department of Chemistry (Canada)], E-mail: kalle@bri.nrc.ca

2002-01-15

A new polymer-stabilized nematic liquid crystal has been characterized for the measurement of biomolecular residual dipolar couplings. Filamentous Pf1 phage were embedded in a polyacrylamide matrix that fixes the orientation of the particles. The alignment was characterized by the quadrupolar splitting of the {sup 2}H NMR water signal and by the measurement of {sup 1}H-{sup 15}N residual dipolar couplings (RDC) in the archeal translation elongation factor 1{beta}. Protein dissolved in the polymer-stabilized medium orients quantitatively as in media without polyacrylamide. We show that the quadrupolar splitting and RDCs are zero in media in which the Pf1 phage particles are aligned at the magic angle. This allows measurement of J and dipolar couplings in a single sample.

Characterization of polyacrylamide-stabilized Pf1 phage liquid crystals for protein NMR spectroscopy

International Nuclear Information System (INIS)

Trempe, Jean-Francois; Morin, Frederick G.; Xia Zhicheng; Marchessault, Robert H.; Gehring, Kalle

2002-01-01

A new polymer-stabilized nematic liquid crystal has been characterized for the measurement of biomolecular residual dipolar couplings. Filamentous Pf1 phage were embedded in a polyacrylamide matrix that fixes the orientation of the particles. The alignment was characterized by the quadrupolar splitting of the 2 H NMR water signal and by the measurement of 1 H- 15 N residual dipolar couplings (RDC) in the archeal translation elongation factor 1β. Protein dissolved in the polymer-stabilized medium orients quantitatively as in media without polyacrylamide. We show that the quadrupolar splitting and RDCs are zero in media in which the Pf1 phage particles are aligned at the magic angle. This allows measurement of J and dipolar couplings in a single sample

Beyond Helper Phage: Using "Helper Cells" to Select Peptide Affinity Ligands.

Directory of Open Access Journals (Sweden)

M Lisa Phipps

Full Text Available Peptides are important affinity ligands for microscopy, biosensing, and targeted delivery. However, because they can have low affinity for their targets, their selection from large naïve libraries can be challenging. When selecting peptidic ligands from display libraries, it is important to: 1 ensure efficient display; 2 maximize the ability to select high affinity ligands; and 3 minimize the effect of the display context on binding. The "helper cell" packaging system has been described as a tool to produce filamentous phage particles based on phagemid constructs with varying display levels, while remaining free of helper phage contamination. Here we report on the first use of this system for peptide display, including the systematic characterization and optimization of helper cells, their inefficient use in antibody display and their use in creating and selecting from a set of phage display peptide libraries. Our libraries were analyzed with unprecedented precision by standard or deep sequencing, and shown to be superior in quality than commercial gold standards. Using our helper cell libraries, we have obtained ligands recognizing Yersinia pestis surface antigen F1V and L-glutamine-binding periplasmic protein QBP. In the latter case, unlike any of the peptide library selections described so far, we used a combination of phage and yeast display to select intriguing peptide ligands. Based on the success of our selections we believe that peptide libraries obtained with helper cells are not only suitable, but preferable to traditional phage display libraries for selection of peptidic ligands.

Phage Therapy in the Era of Synthetic Biology.

Science.gov (United States)

Barbu, E Magda; Cady, Kyle C; Hubby, Bolyn

2016-10-03

For more than a century, bacteriophage (or phage) research has enabled some of the most important discoveries in biological sciences and has equipped scientists with many of the molecular biology tools that have advanced our understanding of replication, maintenance, and expression of genetic material. Phages have also been recognized and exploited as natural antimicrobial agents and nanovectors for gene therapy, but their potential as therapeutics has not been fully exploited in Western medicine because of challenges such as narrow host range, bacterial resistance, and unique pharmacokinetics. However, increasing concern related to the emergence of bacteria resistant to multiple antibiotics has heightened interest in phage therapy and the development of strategies to overcome hurdles associated with bacteriophage therapeutics. Recent progress in sequencing technologies, DNA manipulation, and synthetic biology allowed scientists to refactor the entire bacterial genome of Mycoplasma mycoides, thereby creating the first synthetic cell. These new strategies for engineering genomes may have the potential to accelerate the construction of designer phage genomes with superior therapeutic potential. Here, we discuss the use of phage as therapeutics, as well as how synthetic biology can create bacteriophage with desirable attributes. Copyright © 2016 Cold Spring Harbor Laboratory Press; all rights reserved.

Phage-based magnetoelastic sensor for the detection of Salmonella typhimurium

Science.gov (United States)

Lakshmanan, Ramji S.

In recent years, food-borne illness have garnered the attention of mainstream America with calls now coming from the media for more inspections to ensure the safety of our food supply. Food borne illness from the ingestion of S. typhimurium has been of great concern due to its common occurrence in food products of daily consumption. Annually approximately 80 million cases of food poisoning are reported in the United States alone. The ever growing need for rapid detection of pathogenic microorganisms present in food, environmental and clinical samples has invoked an increased interest in research efforts towards the development of novel diagnostic methodologies. Currently, the detection of bacteria in contaminated food relies on conventional microbiological methods that are time consuming and manpower intensive. This study presents the results of the characterization of a phage-based magnetoelastic biosensor for the detection of Salmonella typhimurium . This affinity-based biosensensor is comprised of a magnetoelastic material as the transducer and filamentous phage as the bio-recognition element. Magnetoelastic materials are ferromagnetic amorphous alloys that change dimensions in the presence of a magnetic field. This effect in combination with the reverse effect (inverse magnetostriction) is utilized in a typical sensor application. A time varying magnetic field causes these sensors to oscillate at a characteristic resonance frequency. The characteristic resonance frequency is dependent on the initial dimensions and physical properties of the material. These materials are of particular interest owing to their unique capability to perform remote (without direct wire contacts to the sensor) sensing, making in-vivo detection and detection in closed containers possible. The phage-immobilized magnetoelastic biosensor was characterized for specificity; dose response in water, spiked apple juice and in spiked milk; selectivity; and longevity. The sensor's sensitivity is

Functional improvement of antibody fragments using a novel phage coat protein III fusion system

DEFF Research Database (Denmark)

Jensen, Kim Bak; Larsen, Martin; Pedersen, Jesper Søndergaard

2002-01-01

Functional expressions of proteins often depend on the presence of host specific factors. Frequently recombinant expression strategies of proteins in foreign hosts, such as bacteria, have been associated with poor yields or significant loss of functionality. Improvements in the performance of het......(s) of the filamentous phage coat protein III. Furthermore, it will be shown that the observed effect is neither due to improved stability nor increased avidity....

Magic-angle spinning NMR of a class I filamentous bacteriophage virus.

Science.gov (United States)

Abramov, Gili; Morag, Omry; Goldbourt, Amir

2011-08-11

The fd bacteriophage is a filamentous virus that is widely used for bio- and nanotechnology applications ranging from phage display to battery materials. The possibility of obtaining a detailed description of its structural properties regardless of its state is therefore essential not only for understanding its physical arrangement and its bacterial infection process but also for many other applications. Here we present a study of the fd phage by magic-angle spinning solid-state NMR. While current structures rely on a Y21M mutant, experiments performed on a strain bearing a wild-type capsid report on high symmetry of the phage and lack of explicit subunit polymorphism. Chemical shift analysis confirmed that the coat protein mostly consists of a rigid right-handed curved α-helix (residues 6-47 of 50), preceded by a flexible loop-structured N-terminus. We were able to qualitatively assign the resonances belonging to the DNA, including the deoxyribose sugars and the thymine bases. These chemical shifts are consistent with base stacking and a C2'-endo/C3'-exo sugar pucker. © 2011 American Chemical Society

Functional display of platelet-binding VWF fragments on filamentous bacteriophage.

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Andrew Yee

Full Text Available von Willebrand factor (VWF tethers platelets to sites of vascular injury via interaction with the platelet surface receptor, GPIb. To further define the VWF sequences required for VWF-platelet interaction, a phage library displaying random VWF protein fragments was screened against formalin-fixed platelets. After 3 rounds of affinity selection, DNA sequencing of platelet-bound clones identified VWF peptides mapping exclusively to the A1 domain. Aligning these sequences defined a minimal, overlapping segment spanning P1254-A1461, which encompasses the C1272-C1458 cystine loop. Analysis of phage carrying a mutated A1 segment (C1272/1458A confirmed the requirement of the cystine loop for optimal binding. Four rounds of affinity maturation of a randomly mutagenized A1 phage library identified 10 and 14 unique mutants associated with enhanced platelet binding in the presence and absence of botrocetin, respectively, with 2 mutants (S1370G and I1372V common to both conditions. These results demonstrate the utility of filamentous phage for studying VWF protein structure-function and identify a minimal, contiguous peptide that bind to formalin-fixed platelets, confirming the importance of the VWF A1 domain with no evidence for another independently platelet-binding segment within VWF. These findings also point to key structural elements within the A1 domain that regulate VWF-platelet adhesion.

Phage annealing proteins promote oligonucleotide-directed mutagenesis in Escherichia coli and mouse ES cells

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Muyrers Joep PP

2003-01-01

Full Text Available Abstract Background The phage protein pairs, RecE/RecT from Rac or Redα/Redβ from λ, initiate efficient double strand break repair (DSBR in Escherichia coli that has proven very useful for DNA engineering. These phage pairs initiate DSBR either by annealing or by another mechanism that is not defined. Results Here we report that these proteins also mediate single strand oligonucleotide repair (ssOR at high efficiencies. The ssOR activity, unlike DSBR, does not require a phage exonuclease (RecE or Redα but only requires a phage annealing protein (RecT or Redβ. Notably, the P22 phage annealing protein Erf, which does not mediate the same DSBR reactions, also delivers ssOR activity. By altering aspects of the oligonucleotides, we document length and design parameters that affect ssOR efficiency to show a simple relationship to homologies either side of the repair site. Notably, ssOR shows strand bias. Oligonucleotides that can prime lagging strand replication deliver more ssOR than their leading complements. This suggests a model in which the annealing proteins hybridize the oligonucleotides to single stranded regions near the replication fork. We also show that ssOR is a highly efficient way to engineer BACs and can be detected in a eukaryotic cell upon expression of a phage annealing protein. Conclusion Phage annealing proteins can initiate the recombination of single stranded oligonucleotides into endogenous targets in Escherichia coli at very high efficiencies. This expands the repertoire of useful DNA engineering strategies, shows promise for applications in eukaryotic cells, and has implications for the unanswered questions regarding DSBR mediated by RecE/RecT and Redα/Redβ.

A Novel Application of Synthetic Biology and Directed Evolution to Engineer Phage-based Antibiotics

Energy Technology Data Exchange (ETDEWEB)

Wu, Meiye [Sandia National Lab. (SNL-CA), Livermore, CA (United States)

2014-09-01

The emergence of multiple drug resistant bacteria poses threats to human health, agriculture and food safety. Annually over 100,000 deaths and up to $20 billion loss to the U.S. economy are attributed to multiple drug resistant bacteria. With only four new chemical antibiotics in the drug development pipeline, we are in dire need of new solutions to address the emerging threat of multiple drug resistance. We propose a paradigm-changing approach to address the multi-drug resistant bacteria problem by utilizing Synthetic Biology (SynBio) methodologies to create and evolve “designer” bacteriophages or phages – viruses that specifically infect bacteria – to infect and kill newly emerging pathogenic bacterial strains WITHOUT the need for chemical antibiotics. A major advantage of using phage to combat pathogenic bacteria is that phages can co-evolve with their bacterial host, and Sandia can be the first in the world to establish an industrial scale Synthetic Biology pipeline for phage directed evolution for safe, targeted, customizable solution to bacterial drug resistance. Since there is no existing phage directed evolution effort within or outside of Sandia, this proposal is suitable as a high-risk LDRD effort to create the first pipeline for such an endeavor. The high potential reward nature of this proposal will be the immediate impact in decontamination and restoration of surfaces and infrastructure, with longer term impact in human or animal therapeutics. The synthetic biology and screening approaches will lead to fundamental knowledge of phage/bacteria co-evolution, making Sandia a world leader in directed evolution of bacteriophages.

The habits of highly effective phages: population dynamics as a framework for identifying therapeutic phages

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James J Bull

2014-11-01

Full Text Available The use of bacteriophages as antibacterial agents is being actively researched on a global scale. Typically, the phages used are isolated from the wild by plating on the bacteria of interest, and a far larger set of candidate phages is often available than can be used in any application. When an excess of phages is available, how should the best phages be identified? Here we consider phage-bacterial population dynamics as a basis for evaluating and predicting phage success. A central question is whether the innate dynamical properties of phages are the determinants of success, or instead, whether extrinsic, indirect effects can be responsible. We address the dynamical perspective, motivated in part by the absence of dynamics in previously suggested principles of phage therapy. Current mathematical models of bacterial-phage dynamics do not capture the realities of in vivo dynamics, nor is this likely to change, but they do give insight to qualitative properties that may be generalizable. In particular, phage adsorption rate may be critical to treatment success, so understanding the effects of the in vivo environment on host availability may allow prediction of useful phages prior to in vivo experimentation. Principles for predicting efficacy may be derived by developing a greater understanding of the in vivo system, or such principles could be determined empirically by comparing phages with known differences in their dynamic properties. The comparative approach promises to be a powerful method of discovering the key to phage success. We offer five recommendations for future study: (i compare phages differing in treatment efficacy to identify the phage properties associated with success, (ii assay dynamics in vivo, (iii understand mechanisms of bacterial escape from phages, (iv test phages in model infections that are relevant to the intended clinical applications, and (v develop new classes of models for phage growth in spatially heterogeneous

Phage transposon mutagenesis.

Science.gov (United States)

Siegrist, M Sloan; Rubin, Eric J

2009-01-01

Phage transduction is an attractive method of genetic manipulation in mycobacteria. PhiMycoMarT7 is well suited for transposon mutagenesis as it is temperature sensitive for replication and contains T7 promoters that promote transcription, a highly active transposase gene, and an Escherichia coli oriR6 K origin of replication. Mycobacterial transposon mutant libraries produced by PhiMycoMarT7 transduction are amenable to both forward and reverse genetic studies. In this protocol, we detail the preparation of PhiMycoMarT7, including a description of the phage, reconstitution of the phage, purification of plaques, preparation of phage stock, and titering of phage stock. We then describe the transduction procedure and finally outline the isolation of individual transposon mutants.

Bacteria, phages and septicemia.

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Ausra Gaidelyte

Full Text Available The use of phages is an attractive option to battle antibiotic resistant bacteria in certain bacterial infections, but the role of phage ecology in bacterial infections is obscure. Here we surveyed the phage ecology in septicemia, the most severe type of bacterial infection. We observed that the majority of the bacterial isolates from septicemia patients spontaneously secreted phages active against other isolates of the same bacterial strain, but not to the strain causing the disease. Such phages were also detected in the initial blood cultures, indicating that phages are circulating in the blood at the onset of sepsis. The fact that most of the septicemic bacterial isolates carry functional prophages suggests an active role of phages in bacterial infections. Apparently, prophages present in sepsis-causing bacterial clones play a role in clonal selection during bacterial invasion.

Morphology of Filamentous Fungi: Linking Cellular Biology to Process Engineering Using Aspergillus niger

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Krull, Rainer; Cordes, Christiana; Horn, Harald; Kampen, Ingo; Kwade, Arno; Neu, Thomas R.; Nörtemann, Bernd

In various biotechnological processes, filamentous fungi, e.g. Aspergillus niger, are widely applied for the production of high value-added products due to their secretion efficiency. There is, however, a tangled relationship between the morphology of these microorganisms, the transport phenomena and the related productivity. The morphological characteristics vary between freely dispersed mycelia and distinct pellets of aggregated biomass. Hence, advantages and disadvantages for mycel or pellet cultivation have to be balanced out carefully. Due to this inadequate understanding of morphogenesis of filamentous microorganisms, fungal morphology, along with reproducibility of inocula of the same quality, is often a bottleneck of productivity in industrial production. To obtain an optimisation of the production process it is of great importance to gain a better understanding of the molecular and cell biology of these microorganisms as well as the approaches in biochemical engineering and particle technique, in particular to characterise the interactions between the growth conditions, cell morphology, spore-hyphae-interactions and product formation. Advances in particle and image analysis techniques as well as micromechanical devices and their applications to fungal cultivations have made available quantitative morphological data on filamentous cells. This chapter provides the ambitious aspects of this line of action, focussing on the control and characterisation of the morphology, the transport gradients and the approaches to understand the metabolism of filamentous fungi. Based on these data, bottlenecks in the morphogenesis of A. niger within the complex production pathways from gene to product should be identified and this may improve the production yield.

Injected phage-displayed-VP28 vaccine reduces shrimp Litopenaeus vannamei mortality by white spot syndrome virus infection.

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Solís-Lucero, G; Manoutcharian, K; Hernández-López, J; Ascencio, F

2016-08-01

White spot syndrome virus (WSSV) is the most important viral pathogen for the global shrimp industry causing mass mortalities with huge economic losses. Recombinant phages are capable of expressing foreign peptides on viral coat surface and act as antigenic peptide carriers bearing a phage-displayed vaccine. In this study, the full-length VP28 protein of WSSV, widely known as potential vaccine against infection in shrimp, was successfully cloned and expressed on M13 filamentous phage. The functionality and efficacy of this vaccine immunogen was demonstrated through immunoassay and in vivo challenge studies. In ELISA assay phage-displayed VP28 was bind to Litopenaeus vannamei immobilized hemocyte in contrast to wild-type M13 phage. Shrimps were injected with 2 × 10(10) cfu animal(-1) single dose of VP28-M13 and M13 once and 48 h later intramuscularly challenged with WSSV to test the efficacy of the vaccine against the infection. All dead challenged shrimps were PCR WSSV-positive. The accumulative mortality of the vaccinated and challenged shrimp groups was significantly lower (36.67%) than the unvaccinated group (66.67%). Individual phenoloxidase and superoxide dismutase activity was assayed on 8 and 48 h post-vaccination. No significant difference was found in those immunological parameters among groups at any sampled time evaluated. For the first time, phage display technology was used to express a recombinant vaccine for shrimp. The highest percentage of relative survival in vaccinated shrimp (RPS = 44.99%) suggest that the recombinant phage can be used successfully to display and deliver VP28 for farmed marine crustaceans. Copyright © 2016 Elsevier Ltd. All rights reserved.

Diversity and Geographical Distribution of Flavobacterium psychrophilum Isolates and Their Phages: Patterns of Susceptibility to Phage Infection and Phage Host Range

DEFF Research Database (Denmark)

Castillo, Daniel; Christiansen, Rói Hammershaimb; Espejo, Romilio

2014-01-01

in disease control requires detailed knowledge about the diversity and dynamics of host susceptibility to phage infection. For this reason, we examined the genetic diversity of 49 F. psychrophilum strains isolated in three different areas (Chile, Denmark, and USA) through direct genome restriction enzyme...... analysis (DGREA) and their susceptibility to 33 bacteriophages isolated in Chile and Denmark, thus covering large geographical (>12,000 km) and temporal (>60 years) scales of isolation. An additional 40 phage-resistant isolates obtained from culture experiments after exposure to specific phages were...... examined for changes in phage susceptibility against the 33 phages. The F. psychrophilum and phage populations isolated from Chile and Denmark clustered into geographically distinct groups with respect to DGREA profile and host range, respectively. However, cross infection between Chilean phage isolates...

Three New Escherichia coli Phages from the Human Gut Show Promising Potential for Phage Therapy.

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Marion Dalmasso

Full Text Available With the emergence of multi-drug resistant bacteria the use of bacteriophages (phages is gaining renewed interest as promising anti-microbial agents. The aim of this study was to isolate and characterize phages from human fecal samples. Three new coliphages, ɸAPCEc01, ɸAPCEc02 and ɸAPCEc03, were isolated. Their phenotypic and genomic characteristics, and lytic activity against biofilm, and in combination with ciprofloxacin, were investigated. All three phages reduced the growth of E. coli strain DPC6051 at multiplicity of infection (MOI between 10-3 and 105. A cocktail of all three phages completely inhibited the growth of E. coli. The phage cocktail also reduced biofilm formation and prevented the emergence of phage-resistant mutants which occurred with single phage. When combined with ciprofloxacin, phage alone or in cocktail inhibited the growth of E. coli and prevented the emergence of resistant mutants. These three new phages are promising biocontrol agents for E. coli infections.

Stumbling across the same phage

DEFF Research Database (Denmark)

Kalatzis, Panagiotis; Rørbo, Nanna Iben; Castillo Bermúdez, Daniel Elías

2017-01-01

46,006 and 54,201 bp. All 19 phages showed high genetic similarity, and 13 phages were genetically identical. Apart from sporadically distributed single nucleotide polymorphisms (SNPs), genetic diversifications were located in three variable regions (VR1, VR2 and VR3) in six of the phage genomes...... was lysogenized by the temperate phages and a genomic analysis of a collection of 31 virulent V. anguillarum showed that the isolated phages were present as prophages in >50% of the strains covering large geographical distances. Further, phage sequences were widely distributed among CRISPR-Cas arrays of publicly...... available sequenced Vibrios. The observed distribution of these specific temperate Vibriophages across large geographical scales may be explained by efficient dispersal of phages and bacteria in the marine environment combined with a mutualistic interaction between temperate phages and their hosts which...

Crystal Structure of the Phage T4 Recombinase UvsX and Its Functional Interaction with the T4 SF2 Helicase UvsW

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Gajewski, Stefan; Webb, Michael R.; Galkin, Vitold; Egelman, Edward H.; Kreuzer, Kenneth N.; White, Stephen W. (Duke); (UV); (SJCH)

2012-07-11

Bacteriophage T4 provides an important model system for studying the mechanism of homologous recombination. We have determined the crystal structure of the T4 UvsX recombinase, and the overall architecture and fold closely resemble those of RecA, including a highly conserved ATP binding site. Based on this new structure, we reanalyzed electron microscopy reconstructions of UvsX-DNA filaments and docked the UvsX crystal structure into two different filament forms: a compressed filament generated in the presence of ADP and an elongated filament generated in the presence of ATP and aluminum fluoride. In these reconstructions, the ATP binding site sits at the protomer interface, as in the RecA filament crystal structure. However, the environment of the ATP binding site is altered in the two filament reconstructions, suggesting that nucleotide cannot be as easily accommodated at the protomer interface of the compressed filament. Finally, we show that the phage helicase UvsW completes the UvsX-promoted strand-exchange reaction, allowing the generation of a simple nicked circular product rather than complex networks of partially exchanged substrates.

The Caulobacter crescentus phage phiCbK: genomics of a canonical phage

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Gill Jason J

2012-10-01

Full Text Available Abstract Background The bacterium Caulobacter crescentus is a popular model for the study of cell cycle regulation and senescence. The large prolate siphophage phiCbK has been an important tool in C. crescentus biology, and has been studied in its own right as a model for viral morphogenesis. Although a system of some interest, to date little genomic information is available on phiCbK or its relatives. Results Five novel phiCbK-like C. crescentus bacteriophages, CcrMagneto, CcrSwift, CcrKarma, CcrRogue and CcrColossus, were isolated from the environment. The genomes of phage phiCbK and these five environmental phage isolates were obtained by 454 pyrosequencing. The phiCbK-like phage genomes range in size from 205 kb encoding 318 proteins (phiCbK to 280 kb encoding 448 proteins (CcrColossus, and were found to contain nonpermuted terminal redundancies of 10 to 17 kb. A novel method of terminal ligation was developed to map genomic termini, which confirmed termini predicted by coverage analysis. This suggests that sequence coverage discontinuities may be useable as predictors of genomic termini in phage genomes. Genomic modules encoding virion morphogenesis, lysis and DNA replication proteins were identified. The phiCbK-like phages were also found to encode a number of intriguing proteins; all contain a clearly T7-like DNA polymerase, and five of the six encode a possible homolog of the C. crescentus cell cycle regulator GcrA, which may allow the phage to alter the host cell’s replicative state. The structural proteome of phage phiCbK was determined, identifying the portal, major and minor capsid proteins, the tail tape measure and possible tail fiber proteins. All six phage genomes are clearly related; phiCbK, CcrMagneto, CcrSwift, CcrKarma and CcrRogue form a group related at the DNA level, while CcrColossus is more diverged but retains significant similarity at the protein level. Conclusions Due to their lack of any apparent relationship to

One-step production of phage-silicon nanoparticles by PLAL as fluorescent nanoprobes for cell identification

Science.gov (United States)

De Plano, Laura M.; Scibilia, Santi; Rizzo, Maria Giovanna; Crea, Sara; Franco, Domenico; Mezzasalma, Angela M.; Guglielmino, Salvatore P. P.

2018-03-01

Silicon nanoparticles (SiNPs) are widely used as promising nanoplatform owing to their high specific surface area, optical properties and biocompatibility. Silicon nanoparticles find possible application in biomedical environment for their potential quantum effects and the functionalization with biomaterials, too. In this work, we propose a new approach for bio-functionalization of SiNPs and M13-engineered bacteriophage, displaying specific peptides that selectively recognize peripheral blood mononuclear cells (PBMC). The "one-step" functionalization is conducted during the laser ablation of silicon plate in buffer solution with engineered bacteriophages, to obtain SiNPs binding bacteriophages (phage-SiNPs). The interaction between SiNPs and bacteriophage is investigated. Particularly, the optical and morphological characterizations of phage-SiNPs are performed by UV-Vis spectroscopy, scanning electron microscopy operating in transmission mode (STEM) and X-ray spectroscopy (EDX). The functionality of phage-SiNPs is investigated through the photoemissive properties in recognition test on PBMC. Our results showed that phage-SiNPs maintain the capability and the activity to bind PBMC within 30 min. The fluorescence of phage-SiNPs allowed to obtain an optical signal on cell type targets. Finally, the proposed strategy demonstrated its potential use in in vitro applications and could be exploited to realize an optical biosensor to detect a specific target.

Phage Therapy: Eco-Physiological Pharmacology

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Stephen T. Abedon

2014-01-01

Full Text Available Bacterial virus use as antibacterial agents, in the guise of what is commonly known as phage therapy, is an inherently physiological, ecological, and also pharmacological process. Physiologically we can consider metabolic properties of phage infections of bacteria and variation in those properties as a function of preexisting bacterial states. In addition, there are patient responses to pathogenesis, patient responses to phage infections of pathogens, and also patient responses to phage virions alone. Ecologically, we can consider phage propagation, densities, distribution (within bodies, impact on body-associated microbiota (as ecological communities, and modification of the functioning of body “ecosystems” more generally. These ecological and physiological components in many ways represent different perspectives on otherwise equivalent phenomena. Comparable to drugs, one also can view phages during phage therapy in pharmacological terms. The relatively unique status of phages within the context of phage therapy as essentially replicating antimicrobials can therefore result in a confluence of perspectives, many of which can be useful towards gaining a better mechanistic appreciation of phage therapy, as I consider here. Pharmacology more generally may be viewed as a discipline that lies at an interface between organism-associated phenomena, as considered by physiology, and environmental interactions as considered by ecology.

Toward Understanding Phage:Host Interactions in the Rumen; Complete Genome Sequences of Lytic Phages Infecting Rumen Bacteria

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Rosalind A. Gilbert

2017-12-01

Full Text Available The rumen is known to harbor dense populations of bacteriophages (phages predicted to be capable of infecting a diverse range of rumen bacteria. While bacterial genome sequencing projects are revealing the presence of phages which can integrate their DNA into the genome of their host to form stable, lysogenic associations, little is known of the genetics of phages which utilize lytic replication. These phages infect and replicate within the host, culminating in host lysis, and the release of progeny phage particles. While lytic phages for rumen bacteria have been previously isolated, their genomes have remained largely uncharacterized. Here we report the first complete genome sequences of lytic phage isolates specifically infecting three genera of rumen bacteria: Bacteroides, Ruminococcus, and Streptococcus. All phages were classified within the viral order Caudovirales and include two phage morphotypes, representative of the Siphoviridae and Podoviridae families. The phage genomes displayed modular organization and conserved viral genes were identified which enabled further classification and determination of closest phage relatives. Co-examination of bacterial host genomes led to the identification of several genes responsible for modulating phage:host interactions, including CRISPR/Cas elements and restriction-modification phage defense systems. These findings provide new genetic information and insights into how lytic phages may interact with bacteria of the rumen microbiome.

Colonisation of a phage susceptible Campylobacter jejuni population in two phage positive broiler flocks.

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Sophie Kittler

Full Text Available The pathogens Campylobacter jejuni and Campylobacter coli are commensals in the poultry intestine and campylobacteriosis is one of the most frequent foodborne diseases in developed and developing countries. Phages were identified to be effective in reducing intestinal Campylobacter load and this was evaluated, in the first field trials which were recently carried out. The aim of this study was to further investigate Campylobacter population dynamics during phage application on a commercial broiler farm. This study determines the superiority in colonisation of a Campylobacter type found in a field trial that was susceptible to phages in in vitro tests. The colonisation factors, i.e. motility and gamma glutamyl transferase activity, were increased in this type. The clustering in phylogenetic comparisons of MALDI-TOF spectra did not match the ST, biochemical phenotype and phage susceptibility. Occurrence of Campylobacter jejuni strains and phage susceptibility types with different colonisation potential seem to play a very important role in the success of phage therapy in commercial broiler houses. Thus, mechanisms of both, phage susceptibility and Campylobacter colonisation should be further investigated and considered when composing phage cocktails.

Phage-Host Interactions in Flavobacterium psychrophilum and the Potential for Phage Therapy in Aquaculture

DEFF Research Database (Denmark)

Christiansen, Rói Hammershaimb

, the increasing problem with antibiotic resistance has led to increased attention to the use of phages for controlling F. psychrophilum infections in aquaculture. In a synopsis and four scientific papers, this PhD project studies the potential and optimizes the use of phage therapy for treatment and prevention......, studies of the genetic diversity and susceptibility patterns of F. psychrophilum strains and phages isolated in three geographically distinct areas (Chile, Denmark, and USA) showed that the strains and phages clustered into geographically distinct groups. However, cross-infectivity between Chilean phage......-phage. In the third paper, a detailed analysis of the resistance mechanisms in F. psychrophilum and six phage resistant mutants was done. The results revealed unique changes in the genomes in all the phage resistant strains and that some of these changes were related to cell surface properties which were suggested...

Novel chitosan film embedded with liposome-encapsulated phage for biocontrol of Escherichia coli O157:H7 in beef.

Science.gov (United States)

Cui, Haiying; Yuan, Lu; Lin, Lin

2017-12-01

In recent years, phages used for the reduction of pathogenic bacteria have fostered many attentions, but they are liable to lost bioactivity in food due to the presence of acidic compounds, enzymes and evaporite materials. To improve the stability of phages, a chitosan edible film containing liposome-encapsulated phage was engineered in the present study. The characteristics of liposome-encapsulated phage and the chitosan film containing liposome-encapsulated phage were investigated. The encapsulation efficiency of phages in liposome reached 57.66±0.12%. Besides, the desirable physical properties of chitosan film were obtained. The chitosan film embedded with liposome-encapsulated phage exhibited high antibacterial activity against Escherichia coli O157:H7, without the impact on the sensory properties of beef. Hence, chitosan film containing liposome-encapsulated phage could be a promising antibacterial packaging for beef preservation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rapid detection of Salmonella typhimurium on fresh spinach leaves using phage-immobilized magnetoelastic biosensors

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Horikawa, Shin; Li, Suiqiong; Chai, Yating; Park, Mi-Kyung; Shen, Wen; Barbaree, James M.; Vodyanoy, Vitaly J.; Chin, Bryan A.

2011-06-01

This paper presents an investigation into the use of magnetoelastic biosensors for the rapid detection of Salmonella typhimurium on fresh spinach leaves. The biosensors used in this investigation were comprised of a strip-shaped, goldcoated sensor platform (2 mm-long) diced from a ferromagnetic, amorphous alloy and a filamentous fd-tet phage which specifically binds with S. typhimurium. After surface blocking with bovine serum albumin, these biosensors were, without any preceding sample preparation, directly placed on wet spinach leaves inoculated with various concentrations of S. typhimurium. Upon contact with cells, the phage binds S. typhimurium to the sensor thereby increasing the total mass of the sensor. This change in mass causes a corresponding decrease in the sensor's resonant frequency. After 25 min, the sensors were collected from the leaf surface and measurements of the resonant frequency were performed immediately. The total assay time was less than 30 min. The frequency changes for measurement sensors (i.e., phageimmobilized) were found to be statistically different from those for control sensors (sensors without phage), down to 5 × 106 cells/ml. The detection limit may be improved by using smaller, micron-sized sensors that will have a higher probability of contacting Salmonella on the rough surfaces of spinach leaves.

Antibody production in response to staphylococcal MS-1 phage cocktail in patients undergoing phage therapy

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Maciej Żaczek

2016-10-01

Full Text Available In this study, we investigated the humoral immune response (through the release of IgG, IgA, and IgM antiphage antibodies to a staphylococcal phage cocktail in patients undergoing experimental phage therapy at the Phage Therapy Unit, Medical Center of the Ludwik Hirszfeld Institute of Immunology and Experimental Therapy in Wrocław, Poland. We also evaluated whether occurring antiphage antibodies had neutralizing properties towards applied phages (K rate. Among 20 examined patients receiving the MS-1 phage cocktail orally and/or locally, the majority did not show a noticeably higher level of antiphage antibodies in their sera during phage administration. Even in those individual cases with an increased immune response, mostly by induction of IgG and IgM, the presence of antiphage antibodies did not translate into unsatisfactory clinical results of phage therapy. On the other hand, a negative outcome of the treatment occurred in some patients who showed relatively weak production of antiphage antibodies before and during treatment. This may imply that possible induction of antiphage antibodies is not an obstacle to the implementation of phage therapy and support our assumption that the outcome of the phage treatment does not primarily depend on the appearance of antiphage antibodies in sera of patients during therapy. These conclusions are in line with our previous findings. The confirmation of this thesis is of great interest as regards the efficacy of phage therapy in humans.

Antibody Production in Response to Staphylococcal MS-1 Phage Cocktail in Patients Undergoing Phage Therapy.

Science.gov (United States)

Żaczek, Maciej; Łusiak-Szelachowska, Marzanna; Jończyk-Matysiak, Ewa; Weber-Dąbrowska, Beata; Międzybrodzki, Ryszard; Owczarek, Barbara; Kopciuch, Agnieszka; Fortuna, Wojciech; Rogóż, Paweł; Górski, Andrzej

2016-01-01

In this study, we investigated the humoral immune response (through the release of IgG, IgA, and IgM antiphage antibodies) to a staphylococcal phage cocktail in patients undergoing experimental phage therapy at the Phage Therapy Unit, Medical Center of the Ludwik Hirszfeld Institute of Immunology and Experimental Therapy in Wrocław, Poland. We also evaluated whether occurring antiphage antibodies had neutralizing properties toward applied phages (K rate). Among 20 examined patients receiving the MS-1 phage cocktail orally and/or locally, the majority did not show a noticeably higher level of antiphage antibodies in their sera during phage administration. Even in those individual cases with an increased immune response, mostly by induction of IgG and IgM, the presence of antiphage antibodies did not translate into unsatisfactory clinical results of phage therapy. On the other hand, a negative outcome of the treatment occurred in some patients who showed relatively weak production of antiphage antibodies before and during treatment. This may imply that possible induction of antiphage antibodies is not an obstacle to the implementation of phage therapy and support our assumption that the outcome of the phage treatment does not primarily depend on the appearance of antiphage antibodies in sera of patients during therapy. These conclusions are in line with our previous findings. The confirmation of this thesis is of great interest as regards the efficacy of phage therapy in humans.

Re-engineering of the PAM1 phage display monoclonal antibody to produce a soluble, versatile anti-homogalacturonan scFv

DEFF Research Database (Denmark)

Manfield, I. W.; Bernal Giraldo, Adriana Jimena; Møller, I.

2006-01-01

Antibody phage display is an increasingly important alternative method for the production of monoclonal antibodies (mAbs) and involves the expression of antibody fragments (scFvs) at the surface of bacteriophage particles. We have previously used this technique to generate a phage mAb (PAM1phage...

Complete genome sequence of Vibrio anguillarum phage CHOED successfully used for phage therapy in aquaculture

DEFF Research Database (Denmark)

Romero, Jaime; Higuera, Gastón; Gajardo, Felipe

2014-01-01

Vibrio anguillarum phage CHOED was isolated from Chilean mussels. It is a virulent phage showing effective inhibition of V. anguillarum. CHOED has potential in phage therapy, because it can protect fish from vibriosis in fish farms. Here, we announce the completely sequenced genome of V....... anguillarum phage CHOED....

Current insights into phage biodiversity and biogeography.

Science.gov (United States)

Thurber, Rebecca Vega

2009-10-01

Phages exert tremendous ecological and evolutionary forces directly on their bacterial hosts. Phage induced cell lysis also indirectly contributes to organic and inorganic nutrient recycling. Phage abundance, diversity, and distribution are therefore important parameters in ecosystem function. The assumption that phage consortia are ubiquitous and homogenous across habitats (everything is everywhere) is currently being re-evaluated. New studies on phage biogeography have found that some phages are globally distributed while others are unique and perhaps endemic to specific environments. Furthermore, advances in technology have allowed scientists to conduct experiments aimed at analyzing phage consortia over temporal scales, and surprisingly have found reoccurring patterns. This review discusses currents in the field of phage ecology with particular focus on efforts to characterize phage diversity and biogeography across various spatial and temporal scales.

HostPhinder: A Phage Host Prediction Tool

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Julia Villarroel

2016-05-01

Full Text Available The current dramatic increase of antibiotic resistant bacteria has revitalised the interest in bacteriophages as alternative antibacterial treatment. Meanwhile, the development of bioinformatics methods for analysing genomic data places high-throughput approaches for phage characterization within reach. Here, we present HostPhinder, a tool aimed at predicting the bacterial host of phages by examining the phage genome sequence. Using a reference database of 2196 phages with known hosts, HostPhinder predicts the host species of a query phage as the host of the most genomically similar reference phages. As a measure of genomic similarity the number of co-occurring k-mers (DNA sequences of length k is used. Using an independent evaluation set, HostPhinder was able to correctly predict host genus and species for 81% and 74% of the phages respectively, giving predictions for more phages than BLAST and significantly outperforming BLAST on phages for which both had predictions. HostPhinder predictions on phage draft genomes from the INTESTI phage cocktail corresponded well with the advertised targets of the cocktail. Our study indicates that for most phages genomic similarity correlates well with related bacterial hosts. HostPhinder is available as an interactive web service [1] and as a stand alone download from the Docker registry [2].

Protein Expression Modifications in Phage-Resistant Mutants of Aeromonas salmonicida after AS-A Phage Treatment

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Catarina Moreirinha

2018-03-01

Full Text Available The occurrence of infections by pathogenic bacteria is one of the main sources of financial loss for the aquaculture industry. This problem often cannot be solved with antibiotic treatment or vaccination. Phage therapy seems to be an alternative environmentally-friendly strategy to control infections. Recognizing the cellular modifications that bacteriophage therapy may cause to the host is essential in order to confirm microbial inactivation, while understanding the mechanisms that drive the development of phage-resistant strains. The aim of this work was to detect cellular modifications that occur after phage AS-A treatment in A. salmonicida, an important fish pathogen. Phage-resistant and susceptible cells were subjected to five successive streak-plating steps and analysed with infrared spectroscopy, a fast and powerful tool for cell study. The spectral differences of both populations were investigated and compared with a phage sensitivity profile, obtained through the spot test and efficiency of plating. Changes in protein associated peaks were found, and these results were corroborated by 1-D electrophoresis of intracellular proteins analysis and by phage sensitivity profiles. Phage AS-A treatment before the first streaking-plate step clearly affected the intracellular proteins expression levels of phage-resistant clones, altering the expression of distinct proteins during the subsequent five successive streak-plating steps, making these clones recover and be phenotypically more similar to the sensitive cells.

Cloning vector

Science.gov (United States)

Guilfoyle, Richard A.; Smith, Lloyd M.

1994-01-01

A vector comprising a filamentous phage sequence containing a first copy of filamentous phage gene X and other sequences necessary for the phage to propagate is disclosed. The vector also contains a second copy of filamentous phage gene X downstream from a promoter capable of promoting transcription in a bacterial host. In a preferred form of the present invention, the filamentous phage is M13 and the vector additionally includes a restriction endonuclease site located in such a manner as to substantially inactivate the second gene X when a DNA sequence is inserted into the restriction site.

Cloning vector

Science.gov (United States)

Guilfoyle, R.A.; Smith, L.M.

1994-12-27

A vector comprising a filamentous phage sequence containing a first copy of filamentous phage gene X and other sequences necessary for the phage to propagate is disclosed. The vector also contains a second copy of filamentous phage gene X downstream from a promoter capable of promoting transcription in a bacterial host. In a preferred form of the present invention, the filamentous phage is M13 and the vector additionally includes a restriction endonuclease site located in such a manner as to substantially inactivate the second gene X when a DNA sequence is inserted into the restriction site. 2 figures.

Phage inactivation by triplet acetone

International Nuclear Information System (INIS)

Gomes, R.A.

1985-01-01

The exposure of lambda phage to triplet acetone is studied. The triplet acetone is obtained from aerobic oxidation of isobutanal catalysed by peroxidase. A decrease of lambda phage ability to infect Escherichia coli is reported, perhaps, partially due to the possible production of lesions in the phage genome. (M.A.C.) [pt

Characterization of Two Virulent Phages of Lactobacillus plantarum

Science.gov (United States)

Briggiler Marcó, Mariángeles; Garneau, Josiane E.; Tremblay, Denise; Quiberoni, Andrea

2012-01-01

We characterized two Lactobacillus plantarum virulent siphophages, ATCC 8014-B1 (B1) and ATCC 8014-B2 (B2), previously isolated from corn silage and anaerobic sewage sludge, respectively. Phage B2 infected two of the eight L. plantarum strains tested, while phage B1 infected three. Phage adsorption was highly variable depending on the strain used. Phage defense systems were found in at least two L. plantarum strains, LMG9211 and WCSF1. The linear double-stranded DNA genome of the pac-type phage B1 had 38,002 bp, a G+C content of 47.6%, and 60 open reading frames (ORFs). Surprisingly, the phage B1 genome has 97% identity with that of Pediococcus damnosus phage clP1 and 77% identity with that of L. plantarum phage JL-1; these phages were isolated from sewage and cucumber fermentation, respectively. The double-stranded DNA (dsDNA) genome of the cos-type phage B2 had 80,618 bp, a G+C content of 36.9%, and 127 ORFs with similarities to those of Bacillus and Lactobacillus strains as well as phages. Some phage B2 genes were similar to ORFs from L. plantarum phage LP65 of the Myoviridae family. Additionally, 6 tRNAs were found in the phage B2 genome. Protein analysis revealed 13 (phage B1) and 9 (phage B2) structural proteins. To our knowledge, this is the first report describing such high identity between phage genomes infecting different genera of lactic acid bacteria. PMID:23042172

Characterizing Phage Genomes for Therapeutic Applications

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Casandra W. Philipson

2018-04-01

Full Text Available Multi-drug resistance is increasing at alarming rates. The efficacy of phage therapy, treating bacterial infections with bacteriophages alone or in combination with traditional antibiotics, has been demonstrated in emergency cases in the United States and in other countries, however remains to be approved for wide-spread use in the US. One limiting factor is a lack of guidelines for assessing the genomic safety of phage candidates. We present the phage characterization workflow used by our team to generate data for submitting phages to the Federal Drug Administration (FDA for authorized use. Essential analysis checkpoints and warnings are detailed for obtaining high-quality genomes, excluding undesirable candidates, rigorously assessing a phage genome for safety and evaluating sequencing contamination. This workflow has been developed in accordance with community standards for high-throughput sequencing of viral genomes as well as principles for ideal phages used for therapy. The feasibility and utility of the pipeline is demonstrated on two new phage genomes that meet all safety criteria. We propose these guidelines as a minimum standard for phages being submitted to the FDA for review as investigational new drug candidates.

Characterizing Phage Genomes for Therapeutic Applications.

Science.gov (United States)

Philipson, Casandra W; Voegtly, Logan J; Lueder, Matthew R; Long, Kyle A; Rice, Gregory K; Frey, Kenneth G; Biswas, Biswajit; Cer, Regina Z; Hamilton, Theron; Bishop-Lilly, Kimberly A

2018-04-10

Multi-drug resistance is increasing at alarming rates. The efficacy of phage therapy, treating bacterial infections with bacteriophages alone or in combination with traditional antibiotics, has been demonstrated in emergency cases in the United States and in other countries, however remains to be approved for wide-spread use in the US. One limiting factor is a lack of guidelines for assessing the genomic safety of phage candidates. We present the phage characterization workflow used by our team to generate data for submitting phages to the Federal Drug Administration (FDA) for authorized use. Essential analysis checkpoints and warnings are detailed for obtaining high-quality genomes, excluding undesirable candidates, rigorously assessing a phage genome for safety and evaluating sequencing contamination. This workflow has been developed in accordance with community standards for high-throughput sequencing of viral genomes as well as principles for ideal phages used for therapy. The feasibility and utility of the pipeline is demonstrated on two new phage genomes that meet all safety criteria. We propose these guidelines as a minimum standard for phages being submitted to the FDA for review as investigational new drug candidates.

Exploration of Phage-Host Interactions in Fish Pathogen Vibrio anguillarum and Anti-Phage Defense Strategies

DEFF Research Database (Denmark)

Tan, Demeng

The disease vibriosis is caused by the bacterial pathogen Vibrio anguillarum and results in large losses in aquaculture both in Denmark and around the world. Antibiotics have been widely used in antimicrobial prophylaxis and treatment of vibriosis. Recently, numerous multidrug-resistant strains...... of V. anguillarum have been isolated, indicating that antibiotic use has to be restricted and alternatives have to be developed. Lytic phages have been demonstrated to play an essential role in preventing bacterial infection. However, phages are also known to play a critical role in the evolution...... of bacterial pathogenicity development. Therefore, successful application of phage therapy in the treatment of vibriosis requires a detailed understanding of phage-host interactions, especially with regards to anti-phage defense mechanisms in the host. Part I. As a first approach, 24 V. anguillarum and 13...

Basics of Antibody Phage Display Technology.

Science.gov (United States)

Ledsgaard, Line; Kilstrup, Mogens; Karatt-Vellatt, Aneesh; McCafferty, John; Laustsen, Andreas H

2018-06-09

Antibody discovery has become increasingly important in almost all areas of modern medicine. Different antibody discovery approaches exist, but one that has gained increasing interest in the field of toxinology and antivenom research is phage display technology. In this review, the lifecycle of the M13 phage and the basics of phage display technology are presented together with important factors influencing the success rates of phage display experiments. Moreover, the pros and cons of different antigen display methods and the use of naïve versus immunized phage display antibody libraries is discussed, and selected examples from the field of antivenom research are highlighted. This review thus provides in-depth knowledge on the principles and use of phage display technology with a special focus on discovery of antibodies that target animal toxins.

MVP: a microbe-phage interaction database.

Science.gov (United States)

Gao, Na L; Zhang, Chengwei; Zhang, Zhanbing; Hu, Songnian; Lercher, Martin J; Zhao, Xing-Ming; Bork, Peer; Liu, Zhi; Chen, Wei-Hua

2018-01-04

Phages invade microbes, accomplish host lysis and are of vital importance in shaping the community structure of environmental microbiota. More importantly, most phages have very specific hosts; they are thus ideal tools to manipulate environmental microbiota at species-resolution. The main purpose of MVP (Microbe Versus Phage) is to provide a comprehensive catalog of phage-microbe interactions and assist users to select phage(s) that can target (and potentially to manipulate) specific microbes of interest. We first collected 50 782 viral sequences from various sources and clustered them into 33 097 unique viral clusters based on sequence similarity. We then identified 26 572 interactions between 18 608 viral clusters and 9245 prokaryotes (i.e. bacteria and archaea); we established these interactions based on 30 321 evidence entries that we collected from published datasets, public databases and re-analysis of genomic and metagenomic sequences. Based on these interactions, we calculated the host range for each of the phage clusters and accordingly grouped them into subgroups such as 'species-', 'genus-' and 'family-' specific phage clusters. MVP is equipped with a modern, responsive and intuitive interface, and is freely available at: http://mvp.medgenius.info. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Engineered enzymatically active bacteriophages and methods of uses thereof

Energy Technology Data Exchange (ETDEWEB)

Collins, James J [Newton, MA; Kobayashi, Hideki [Yokohama, JP; Kearn, Mads [Ottawa, CA; Araki, Michihiro [Minatoku, JP; Friedland, Ari [Boston, MA; Lu, Timothy Kuan-Ta [Palo Alto, CA

2012-05-22

The present invention provides engineered bacteriophages that express at least one biofilm degrading enzyme on their surface and uses thereof for degrading bacterial biofilms. The invention also provides genetically engineered bacteriophages expressing the biofilm degrading enzymes and proteins necessary for the phage to replicate in different naturally occurring biofilm producing bacteria. The phages of the invention allow a method of biofilm degradation by the use of one or only a few administration of the phage because the system using these phages is self perpetuating, and capable of degrading biofilm even when the concentration of bacteria within the biofilm is low.

Characterization of the integrated filamentous phage Pf5 and its involvement in small-colony formation

NARCIS (Netherlands)

Mooij, Marlies J.; Drenkard, Eliana; Llamas, María A.; Vandenbroucke-Grauls, Christina M. J. E.; Savelkoul, Paul H. M.; Ausubel, Frederick M.; Bitter, Wilbert

2007-01-01

Bacteriophages play an important role in bacterial virulence and phenotypic variation. It has been shown that filamentous bacteriophage Pf4 of Pseudomonas aeruginosa strain PAO1 mediates the formation of small-colony variants (SCVs) in biofilms. This morphology type is associated with parameters of

Control of Pierce's Disease by Phage.

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Mayukh Das

Full Text Available Pierce's Disease (PD of grapevines, caused by Xylella fastidiosa subsp. fastidiosa (Xf, is a limiting factor in the cultivation of grapevines in the US. There are presently no effective control methods to prevent or treat PD. The therapeutic and prophylactic efficacy of a phage cocktail composed of four virulent (lytic phages was evaluated for control of PD. Xf levels in grapevines were significantly reduced in therapeutically or prophylactically treated grapevines. PD symptoms ceased to progress one week post-therapeutic treatment and symptoms were not observed in prophylactically treated grapevines. Cocktail phage levels increased in grapevines in the presence of the host. No in planta phage-resistant Xf isolates were obtained. Moreover, Xf mutants selected for phage resistance in vitro did not cause PD symptoms. Our results indicate that phages have great potential for biocontrol of PD and other economically important diseases caused by Xylella.

Phase variable expression of a single phage receptor in Campylobacter jejuni NCTC12662 influences sensitivity toward several diverse CPS-dependent phages

DEFF Research Database (Denmark)

Gencay, Yilmaz Emre; Sørensen, Martine C.H.; Wenzel, Cory Q.

2018-01-01

Campylobacter jejuni NCTC12662 is sensitive to infection by many Campylobacter bacteriophages. Here we used this strain to investigate the molecular mechanism behind phage resistance development when exposed to a single phage and demonstrate how phase variable expression of one surface component...... influences phage sensitivity against many diverse C. jejuni phages. When C. jejuni NCTC12662 was exposed to phage F207 overnight, 25% of the bacterial cells were able to grow on a lawn of phage F207, suggesting that resistance develops at a high frequency. One resistant variant, 12662R, was further...... characterized and shown to be an adsorption mutant. Plaque assays using our large phage collection showed that seven out of 36 diverse capsular polysaccharide (CPS)-dependent phages could not infect 12662R, whereas the remaining phages formed plaques on 12662R with reduced efficiencies. Analysis of the CPS...

Phages in the Human Body.

Science.gov (United States)

Navarro, Ferran; Muniesa, Maite

2017-01-01

Bacteriophages, viruses that infect bacteria, have re-emerged as powerful regulators of bacterial populations in natural ecosystems. Phages invade the human body, just as they do other natural environments, to such an extent that they are the most numerous group in the human virome. This was only revealed in recent metagenomic studies, despite the fact that the presence of phages in the human body was reported decades ago. The influence of the presence of phages in humans has yet to be evaluated; but as in marine environments, a clear role in the regulation of bacterial populations could be envisaged, that might have an impact on human health. Moreover, phages are excellent vehicles of genetic transfer, and they contribute to the evolution of bacterial cells in the human body by spreading and acquiring DNA horizontally. The abundance of phages in the human body does not pass unnoticed and the immune system reacts to them, although it is not clear to what extent. Finally, the presence of phages in human samples, which most of the time is not considered, can influence and bias microbiological and molecular results; and, in view of the evidences, some studies suggest that more attention needs to be paid to their interference.

Current taxonomy of phages infecting lactic acid bacteria

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Jennifer eMahony

2014-01-01

Full Text Available Phages infecting lactic acid bacteria have been the focus of significant research attention over the past three decades. Through the isolation and characterization of hundreds of phage isolates, it has been possible to classify phages of the dairy starter and adjunct bacteria Lactococus lactis, Streptococcus thermophilus, Leuconostoc spp. and Lactobacillus spp. Among these, phages of L. lactis have been most thoroughly scrutinized and serve as an excellent model system to address issues that arise when attempting taxonomic classification of phages infecting other LAB species. Here, we present an overview of the current taxonomy of phages infecting LAB genera of industrial significance, the methods employed in these taxonomic efforts and how these may be employed for the taxonomy of phages of currently underrepresented and emerging phage species.

Formulation, stabilisation and encapsulation of bacteriophage for phage therapy.

Science.gov (United States)

Malik, Danish J; Sokolov, Ilya J; Vinner, Gurinder K; Mancuso, Francesco; Cinquerrui, Salvatore; Vladisavljevic, Goran T; Clokie, Martha R J; Garton, Natalie J; Stapley, Andrew G F; Kirpichnikova, Anna

2017-11-01

Against a backdrop of global antibiotic resistance and increasing awareness of the importance of the human microbiota, there has been resurgent interest in the potential use of bacteriophages for therapeutic purposes, known as phage therapy. A number of phage therapy phase I and II clinical trials have concluded, and shown phages don't present significant adverse safety concerns. These clinical trials used simple phage suspensions without any formulation and phage stability was of secondary concern. Phages have a limited stability in solution, and undergo a significant drop in phage titre during processing and storage which is unacceptable if phages are to become regulated pharmaceuticals, where stable dosage and well defined pharmacokinetics and pharmacodynamics are de rigueur. Animal studies have shown that the efficacy of phage therapy outcomes depend on the phage concentration (i.e. the dose) delivered at the site of infection, and their ability to target and kill bacteria, arresting bacterial growth and clearing the infection. In addition, in vitro and animal studies have shown the importance of using phage cocktails rather than single phage preparations to achieve better therapy outcomes. The in vivo reduction of phage concentration due to interactions with host antibodies or other clearance mechanisms may necessitate repeated dosing of phages, or sustained release approaches. Modelling of phage-bacterium population dynamics reinforces these points. Surprisingly little attention has been devoted to the effect of formulation on phage therapy outcomes, given the need for phage cocktails, where each phage within a cocktail may require significantly different formulation to retain a high enough infective dose. This review firstly looks at the clinical needs and challenges (informed through a review of key animal studies evaluating phage therapy) associated with treatment of acute and chronic infections and the drivers for phage encapsulation. An important driver

Viruses Infecting a Freshwater Filamentous Cyanobacterium (Nostoc sp. Encode a Functional CRISPR Array and a Proteobacterial DNA Polymerase B

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Caroline Chénard

2016-06-01

Full Text Available Here we present the first genomic characterization of viruses infecting Nostoc, a genus of ecologically important cyanobacteria that are widespread in freshwater. Cyanophages A-1 and N-1 were isolated in the 1970s and infect Nostoc sp. strain PCC 7210 but remained genomically uncharacterized. Their 68,304- and 64,960-bp genomes are strikingly different from those of other sequenced cyanophages. Many putative genes that code for proteins with known functions are similar to those found in filamentous cyanobacteria, showing a long evolutionary history in their host. Cyanophage N-1 encodes a CRISPR array that is transcribed during infection and is similar to the DR5 family of CRISPRs commonly found in cyanobacteria. The presence of a host-related CRISPR array in a cyanophage suggests that the phage can transfer the CRISPR among related cyanobacteria and thereby provide resistance to infection with competing phages. Both viruses also encode a distinct DNA polymerase B that is closely related to those found in plasmids of Cyanothece sp. strain PCC 7424, Nostoc sp. strain PCC 7120, and Anabaena variabilis ATCC 29413. These polymerases form a distinct evolutionary group that is more closely related to DNA polymerases of proteobacteria than to those of other viruses. This suggests that the polymerase was acquired from a proteobacterium by an ancestral virus and transferred to the cyanobacterial plasmid. Many other open reading frames are similar to a prophage-like element in the genome of Nostoc sp. strain PCC 7524. The Nostoc cyanophages reveal a history of gene transfers between filamentous cyanobacteria and their viruses that have helped to forge the evolutionary trajectory of this previously unrecognized group of phages.

Vibrio vulnificus phage PV94 is closely related to temperate phages of V. cholerae and other Vibrio species.

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Mark Pryshliak

Full Text Available BACKGROUND: Vibrio vulnificus is an important pathogen which can cause serious infections in humans. Yet, there is limited knowledge on its virulence factors and the question whether temperate phages might be involved in pathogenicity, as is the case with V. cholerae. Thus far, only two phages (SSP002 and VvAW1 infecting V. vulnificus have been genetically characterized. These phages were isolated from the environment and are not related to Vibrio cholerae phages. The lack of information on temperate V. vulnificus phages prompted us to isolate those phages from lysogenic strains and to compare them with phages of other Vibrio species. RESULTS: In this study the temperate phage PV94 was isolated from a V. vulnificus biotype 1 strain by mitomycin C induction. PV94 is a myovirus whose genome is a linear double-stranded DNA of 33,828 bp with 5'-protruding ends. Sequence analysis of PV94 revealed a modular organization of the genome. The left half of the genome comprising the immunity region and genes for the integrase, terminase and replication proteins shows similarites to V. cholerae kappa phages whereas the right half containing genes for structural proteins is closely related to a prophage residing in V. furnissii NCTC 11218. CONCLUSION: We present the first genomic sequence of a temperate phage isolated from a human V. vulnificus isolate. The sequence analysis of the PV94 genome demonstrates the wide distribution of closely related prophages in various Vibrio species. Moreover, the mosaicism of the PV94 genome indicates a high degree of horizontal genetic exchange within the genus Vibrio, by which V. vulnificus might acquire virulence-associated genes from other species.

Large palindromes in the lambda phage genome are preserved in a rec/sup +/ host by inhibiting lambda DNA replication

Energy Technology Data Exchange (ETDEWEB)

Shurvinton, C.E.; Stahl, M.M.; Stahl, F.W.

1987-03-01

A large palindrome carried by phage lambda has been shown to prevent growth of the phage on a rec/sup +/ strain of Escherichia coli. The phage do form plaques on recBC sbcB strains, but the palindrome is not stable - deletions that either destroy the palindrome or diminish its size overgrow the original engineered palindrome-containing phage. The authors have prepared stocks of lambda carrying a palindrome that is 2 x 4200 base pairs long. lambda phage were density labeled by UV induction of lysogens grown in minimal medium containing (/sup 13/C) glucose and /sup 15/NH/sub 4/Cl. These phage stocks are produced by induction of a lysogen in which the two halves of the palindrome are stored at opposite ends of the prophage and are of sufficient titer (10/sup 9/ phage per ml) to enable one-step growth experiments with replication-blocked phage. They find that the large palindrome as well as a lesser palindrome of 2 x 265 base pairs are recovered intact among particles carrying unreplicated chromosomes following such an infection of a rec/sup +/ host. they propose that DNA replication drives the extrusion of palindromic sequences in vivo, forming secondary structures that are substrates for the recBC and sbcB gene products.

Phages of life - the path to pharma.

Science.gov (United States)

Forde, Amanda; Hill, Colin

2018-02-01

Bacteriophage (phage) therapy has encountered both enthusiasm and scepticism in the past century. New antimicrobial strategies against lethal pathogens are now a top priority for the World Health Organization, and although compassionate use of phages recently met with significant success, regulated clinical interventions seem unlikely in the near future. The hundredth anniversary of their discovery seems an appropriate time for a revival of phage therapy, particularly as the dilemma of antibiotic resistance grows. Phages are ubiquitous in the environment, on our food and in and on our bodies. Their influence on human health is currently being evaluated, and in this mini-review, we examine data from recent metagenomic studies that propose a role for phages in the structure of the microbiome and in health and disease. We assess evidence for phages as vehicles for gene transfer in the context of antibiotic resistance and discuss challenges and opportunities along the critical path from phage discovery to a patient-focused pharmaceutical intervention. © 2017 The British Pharmacological Society.

Heat tolerance of dairy lactococcal c2 phages

DEFF Research Database (Denmark)

Nielsen, Cecilie Lykke Marvig; Basheer, Aideh; Neve, H.

2011-01-01

-order kinetics with correlation coefficients of 0.96–0.99. D70-values of 12 s and 16.6 min were calculated for the most sensitive and resistant phage, respectively. Release of phage DNA from capsids, and disintegration of phage heads and tails were among the first morphological changes observed for moderately...... thermal inactivated lysates (15% phage inactivation) of the heat tolerant phage P635....

Efficient engineering of a bacteriophage genome using the type I-E CRISPR-Cas system.

Science.gov (United States)

Kiro, Ruth; Shitrit, Dror; Qimron, Udi

2014-01-01

The clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) system has recently been used to engineer genomes of various organisms, but surprisingly, not those of bacteriophages (phages). Here we present a method to genetically engineer the Escherichia coli phage T7 using the type I-E CRISPR-Cas system. T7 phage genome is edited by homologous recombination with a DNA sequence flanked by sequences homologous to the desired location. Non-edited genomes are targeted by the CRISPR-Cas system, thus enabling isolation of the desired recombinant phages. This method broadens CRISPR Cas-based editing to phages and uses a CRISPR-Cas type other than type II. The method may be adjusted to genetically engineer any bacteriophage genome.

The Staphylococci Phages Family: An Overview

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Laurence Van Melderen

2012-11-01

Full Text Available Due to their crucial role in pathogenesis and virulence, phages of Staphylococcus aureus have been extensively studied. Most of them encode and disseminate potent staphylococcal virulence factors. In addition, their movements contribute to the extraordinary versatility and adaptability of this prominent pathogen by improving genome plasticity. In addition to S. aureus, phages from coagulase-negative Staphylococci (CoNS are gaining increasing interest. Some of these species, such as S. epidermidis, cause nosocomial infections and are therefore problematic for public health. This review provides an overview of the staphylococcal phages family extended to CoNS phages. At the morphological level, all these phages characterized so far belong to the Caudovirales order and are mainly temperate Siphoviridae. At the molecular level, comparative genomics revealed an extensive mosaicism, with genes organized into functional modules that are frequently exchanged between phages. Evolutionary relationships within this family, as well as with other families, have been highlighted. All these aspects are of crucial importance for our understanding of evolution and emergence of pathogens among bacterial species such as Staphylococci.

Development of a phage typing system for Staphylococcus hyicus

DEFF Research Database (Denmark)

Wegener, Henrik Caspar

1993-01-01

Bacteriophages were released by 98% of 100 Staphylococcus hyicus strains studied after treatment with mitomycin C. Twenty-three phages with different lytic spectra were included in a phage typing system and used f or typing S. hyicus. On a test-set of 100 epidemiologically unrelated S. hyicus...... strains isolated from Danish pig herds, the phages were able to type 92% of the strains, producing 16 different phage types. Reproducibility of the phage typing system after subculture of the strains and using fresh phage stock was 96%. Typability ranged from 52 to 80% when typing porcine strains...... originating from other countries. Although phages were isolated from porcine skin strains exclusively, the system produced phage types in S. hyicus strains of bovine origin. Ten strains of S. aureus and S. chromogenes were not typable by these phages. Strains belonging to one phage type (A/B/C/W) were...

Sensitive detection of viable Escherichia coli O157:H7 from foods using a luciferase-reporter phage phiV10lux.

Science.gov (United States)

Kim, Jinwoo; Kim, Minsik; Kim, Seongmi; Ryu, Sangryeol

2017-08-02

Escherichia coli O157:H7, a major foodborne pathogen, is a major public health concern associated with life-threatening diseases such as hemolytic uremic syndrome. To alleviate this burden, a sensitive and rapid system is required to detect this pathogen in various kinds of foods. Herein, we propose a phage-based pathogen detection method to replace laborious and time-consuming conventional methods. We engineered an E. coli O157:H7-specific phage phiV10 to rapidly and sensitively detect this notorious pathogen. The luxCDABE operon was introduced into the phiV10 genome and allowed the engineered phage phiV10lux to generate bioluminescence proportional to the number of viable E. coli O157:H7 cells without any substrate addition. The phage phiV10lux was able to detect at least 1CFU/ml of E. coli O157:H7 in a pure culture within 40min after 5h of pre-incubation. In artificially contaminated romaine lettuce, apple juice (pH3.51), and ground beef, the reporter phage could detect approximately 10CFU/cm 2 , 13CFU/ml, and 17CFU/g of E. coli O157:H7, respectively. Taken together, the constructed reporter phage phiV10lux could be applied as a powerful tool for rapid and sensitive detection of live E. coli O157:H7 in foods. Copyright © 2017 Elsevier B.V. All rights reserved.

A broadly implementable research course in phage discovery and genomics for first-year undergraduate students.

Science.gov (United States)

Jordan, Tuajuanda C; Burnett, Sandra H; Carson, Susan; Caruso, Steven M; Clase, Kari; DeJong, Randall J; Dennehy, John J; Denver, Dee R; Dunbar, David; Elgin, Sarah C R; Findley, Ann M; Gissendanner, Chris R; Golebiewska, Urszula P; Guild, Nancy; Hartzog, Grant A; Grillo, Wendy H; Hollowell, Gail P; Hughes, Lee E; Johnson, Allison; King, Rodney A; Lewis, Lynn O; Li, Wei; Rosenzweig, Frank; Rubin, Michael R; Saha, Margaret S; Sandoz, James; Shaffer, Christopher D; Taylor, Barbara; Temple, Louise; Vazquez, Edwin; Ware, Vassie C; Barker, Lucia P; Bradley, Kevin W; Jacobs-Sera, Deborah; Pope, Welkin H; Russell, Daniel A; Cresawn, Steven G; Lopatto, David; Bailey, Cheryl P; Hatfull, Graham F

2014-02-04

Engaging large numbers of undergraduates in authentic scientific discovery is desirable but difficult to achieve. We have developed a general model in which faculty and teaching assistants from diverse academic institutions are trained to teach a research course for first-year undergraduate students focused on bacteriophage discovery and genomics. The course is situated within a broader scientific context aimed at understanding viral diversity, such that faculty and students are collaborators with established researchers in the field. The Howard Hughes Medical Institute (HHMI) Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) course has been widely implemented and has been taken by over 4,800 students at 73 institutions. We show here that this alliance-sourced model not only substantially advances the field of phage genomics but also stimulates students' interest in science, positively influences academic achievement, and enhances persistence in science, technology, engineering, and mathematics (STEM) disciplines. Broad application of this model by integrating other research areas with large numbers of early-career undergraduate students has the potential to be transformative in science education and research training. Engagement of undergraduate students in scientific research at early stages in their careers presents an opportunity to excite students about science, technology, engineering, and mathematics (STEM) disciplines and promote continued interests in these areas. Many excellent course-based undergraduate research experiences have been developed, but scaling these to a broader impact with larger numbers of students is challenging. The Howard Hughes Medical Institute (HHMI) Science Education Alliance Phage Hunting Advancing Genomics and Evolutionary Science (SEA-PHAGES) program takes advantage of the huge size and diversity of the bacteriophage population to engage students in discovery of new viruses, genome

[Isolation and characterization of siphovirus phages infecting bovine Streptococcus agalactiae].

Science.gov (United States)

Bai, Qinqin; Yang, Yongchun; Lu, Chengping

2016-02-04

To isolate and identify Streptococcus agalactiae phages and screen candidate phages to control infection caused by bovine S. agalactiae. We used two methods for isolation of S. agalactiae phages, namely (1) isolation of phages from milk and environmental samples, and (2) isolation of phages via induction of lysogens with Mitomycin C. Double-layer agar culture method was used to purify phages. Then the newly obtained phages, with S. agalactiae phage JX01 isolated from mastitis milk, were comparatively analyzed in the following aspects: morphology of phages by transmission electron microscopy, host range of phages to 55 S. agalactiae strains and other Streptococcus strains, phages DNA using EcoR I, Xba I, Pst I and Sal I, the optical multiplicity of infection, absorption curve and one step growth curve, and the stability of phages at different storage conditions. The comparative analysis of the 3 novel phages LYGO9, HZ04 and pA11 (induced from S. agalctiae bovine clinical isolate HAJL2011070601) with JX01 showed that the 4 phages were classified as the member of Siphovirdae family. EcoR I, Sal I, Xba I and Pst I separately digested the 4 phages DNA provided 4, 3, 3 and 2 profiles, respectively. This suggested that they were different strains. All the 4 phages specifically infected bovine S. agalactiae isolates. LYGO9, pA11, JX01 and HZ04 could lyse 12, 13, 20 and 23 of 42 tested bovine S. agalctiae isolates, respectively. This clearly indicated that these 4 phages are closely related. The 3 new phages which specifically lyse bovine S. agalactiae isolates are siphovirus phages. Phage LYGO9 was shown having a short latent period and a larger burst size.

Phage-Bacterial Dynamics with Spatial Structure: Self Organization around Phage Sinks Can Promote Increased Cell Densities.

Science.gov (United States)

Bull, James J; Christensen, Kelly A; Scott, Carly; Jack, Benjamin R; Crandall, Cameron J; Krone, Stephen M

2018-01-29

Bacteria growing on surfaces appear to be profoundly more resistant to control by lytic bacteriophages than do the same cells grown in liquid. Here, we use simulation models to investigate whether spatial structure per se can account for this increased cell density in the presence of phages. A measure is derived for comparing cell densities between growth in spatially structured environments versus well mixed environments (known as mass action). Maintenance of sensitive cells requires some form of phage death; we invoke death mechanisms that are spatially fixed, as if produced by cells. Spatially structured phage death provides cells with a means of protection that can boost cell densities an order of magnitude above that attained under mass action, although the effect is sometimes in the opposite direction. Phage and bacteria self organize into separate refuges, and spatial structure operates so that the phage progeny from a single burst do not have independent fates (as they do with mass action). Phage incur a high loss when invading protected areas that have high cell densities, resulting in greater protection for the cells. By the same metric, mass action dynamics either show no sustained bacterial elevation or oscillate between states of low and high cell densities and an elevated average. The elevated cell densities observed in models with spatial structure do not approach the empirically observed increased density of cells in structured environments with phages (which can be many orders of magnitude), so the empirical phenomenon likely requires additional mechanisms than those analyzed here.

Binding of phage displayed Bacillus subtilis lipase A to a phosphonate suicide inhibitor

NARCIS (Netherlands)

Dröge, M.J; Ruggeberg, C.J.; van der Sloot, Almer Martinus; Schimmel, J.; Dijkstra, Durk; Verhaert, R.M D; Reetz, M.T.; Quax, Wim; Droge, MJ; Dijkstra, DS

2003-01-01

Phage display can be used as a protein engineering tool to select proteins with desirable binding properties from a library of randomly constructed mutants. Here, we describe the development of this method for the directed evolution of Bacillus subtilis lipase A, an enzyme that has marked properties

Phage Therapy -- Everything Old Is New again

Directory of Open Access Journals (Sweden)

Andrew M Kropinski

2006-01-01

Full Text Available The study of bacterial viruses (bacteriophages or phages proved pivotal in the nascence of the disciplines of molecular biology and microbial genetics, providing important information on the central processes of the bacterial cell (DNA replication, transcription and translation and on how DNA can be transferred from one cell to another. As a result of the pioneering genetics studies and modern genomics, it is now known that phages have contributed to the evolution of the microbial cell and to its pathogenic potential. Because of their ability to transmit genes, phages have been exploited to develop cloning vector systems. They also provide a plethora of enzymes for the modern molecular biologist. Until the introduction of antibiotics, phages were used to treat bacterial infections (with variable success. Western science is now having to re-evaluate the application of phage therapy -- a therapeutic modality that never went out of vogue in Eastern Europe -- because of the emergence of an alarming number of antibiotic-resistant bacteria. The present article introduces the reader to phage biology, and the benefits and pitfalls of phage therapy in humans and animals.

Phage-mediated counting by the naked eye of miRNA molecules at attomolar concentrations in a Petri dish

Science.gov (United States)

Zhou, Xin; Cao, Peng; Zhu, Ye; Lu, Wuguang; Gu, Ning; Mao, Chuanbin

2015-10-01

The ability to count biomolecules such as cancer-biomarker miRNAs with the naked eye is seemingly impossible in molecular diagnostics. Here, we show an ultrasensitive naked-eye-counting strategy for quantifying miRNAs by employing T7 phage--a bacteria-specific virus nanoparticle--as a surrogate. The phage is genetically engineered to become fluorescent and capable of binding a miRNA-capturing gold nanoparticle (GNP) in a one-to-one manner. Target miRNAs crosslink the resultant phage-GNP couple and miRNA-capturing magnetic microparticles, forming a sandwich complex containing equimolar phage and miRNA. The phage is then released from the complex and developed into one macroscopic fluorescent plaque in a Petri dish by plating it in a host bacterial medium. Counting the plaques by the naked eye enables the quantification of miRNAs with detection limits of ~3 and ~5 aM for single-target and two-target miRNAs, respectively. This approach offers ultrasensitive and convenient quantification of disease biomarkers by the naked eye.

Isolation and Characterization of Phages Infecting Bacillus subtilis

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Anna Krasowska

2015-01-01

Full Text Available Bacteriophages have been suggested as an alternative approach to reduce the amount of pathogens in various applications. Bacteriophages of various specificity and virulence were isolated as a means of controlling food-borne pathogens. We studied the interaction of bacteriophages with Bacillus species, which are very often persistent in industrial applications such as food production due to their antibiotic resistance and spore formation. A comparative study using electron microscopy, PFGE, and SDS-PAGE as well as determination of host range, pH and temperature resistance, adsorption rate, latent time, and phage burst size was performed on three phages of the Myoviridae family and one phage of the Siphoviridae family which infected Bacillus subtilis strains. The phages are morphologically different and characterized by icosahedral heads and contractile (SIOΦ, SUBω, and SPOσ phages or noncontractile (ARπ phage tails. The genomes of SIOΦ and SUBω are composed of 154 kb. The capsid of SIOΦ is composed of four proteins. Bacteriophages SPOσ and ARπ have genome sizes of 25 kbp and 40 kbp, respectively. Both phages as well as SUBω phage have 14 proteins in their capsids. Phages SIOΦ and SPOσ are resistant to high temperatures and to the acid (4.0 and alkaline (9.0 and 10.0 pH.

Pseudomonas predators: understanding and exploiting phage-host interactions.

Science.gov (United States)

De Smet, Jeroen; Hendrix, Hanne; Blasdel, Bob G; Danis-Wlodarczyk, Katarzyna; Lavigne, Rob

2017-09-01

Species in the genus Pseudomonas thrive in a diverse set of ecological niches and include crucial pathogens, such as the human pathogen Pseudomonas aeruginosa and the plant pathogen Pseudomonas syringae. The bacteriophages that infect Pseudomonas spp. mirror the widespread and diverse nature of their hosts. Therefore, Pseudomonas spp. and their phages are an ideal system to study the molecular mechanisms that govern virus-host interactions. Furthermore, phages are principal catalysts of host evolution and diversity, which directly affects the ecological roles of environmental and pathogenic Pseudomonas spp. Understanding these interactions not only provides novel insights into phage biology but also advances the development of phage therapy, phage-derived antimicrobial strategies and innovative biotechnological tools that may be derived from phage-bacteria interactions.

Simultaneous display of two large proteins on the head and tail of bacteriophage lambda

Science.gov (United States)

2013-01-01

Background Consistent progress in the development of bacteriophage lambda display platform as an alternative to filamentous phage display system was achieved in the recent years. The lambda phage has been engineered to display efficiently multiple copies of peptides or even large protein domains providing a powerful tool for screening libraries of peptides, proteins and cDNA. Results In the present work we describe an original method for dual display of large proteins on the surface of lambda particles. An anti-CEA single-chain antibody fragment and green fluorescent protein or alkaline phosphatase were simultaneously displayed by engineering both gpD and gpV lambda proteins. Conclusions Here we show that such modified phage particles can be used for the detection of target molecules in vitro and in vivo. Dual expression of functional moieties on the surface of the lambda phage might open the way to generation of a new class of diagnostic and therapeutic targeted nanoparticles. PMID:24073829

Phage typing of Staphylococcus saprophyticus.

Science.gov (United States)

Torres Pereira, A.; Melo Cristino, J. A.

1991-01-01

This study included 502 staphylococcus strains; Staphylococcus saprophyticus (297 strains) S. cohnii (47), S. xylosus (10), S. epidermidis (67) and S. aureus (81). Mitomycin C induction was performed on 100 isolates of S. saprophyticus and all induced strains were reacted with each other. Twenty-six strains proved to be lysogenic. Phages were propagated and titrated. With 12 of the phages there were three frequent associations, named lytic groups A, B and C, which included 75% of all typable strains. Typability of the system was 45% and reproducibility was between 94.2% and 100%. Phages did not lyse S. aureus and S. epidermidis strains, but they lysed S. saprophyticus and only rare strains of other novobiocin resistant species. Effective S. saprophyticus typing serves ecological purposes and tracing the origin of urinary strains from the skin or mucous membranes. Phage typing in association with plasmid profiling previously described, are anticipated as complementary methods with strong discriminatory power for differentiating among S. saprophyticus strains. PMID:1752305

Filament heater current modulation for increased filament lifetime

International Nuclear Information System (INIS)

Paul, J.D.; Williams, H.E. III.

1996-01-01

The surface conversion H-minus ion source employs two 60 mil tungsten filaments which are approximately 17 centimeters in length. These filaments are heated to approximately 2,800 degrees centigrade by 95--100 amperes of DC heater current. The arc is struck at a 120 hertz rate, for 800 microseconds and is generally run at 30 amperes peak current. Although sputtering is considered a contributing factor in the demise of the filament, evaporation is of greater concern. If the peak arc current can be maintained with less average heater current, the filament evaporation rate for this arc current will diminish. In the vacuum of an ion source, the authors expect the filaments to retain much of their heat throughout a 1 millisecond (12% duty) loss of heater current. A circuit to eliminate 100 ampere heater currents from filaments during the arc pulse was developed. The magnetic field due to the 100 ampere current tends to hold electrons to the filament, decreasing the arc current. By eliminating this magnetic field, the arc should be more efficient, allowing the filaments to run at a lower average heater current. This should extend the filament lifetime. The circuit development and preliminary filament results are discussed

YpfΦ : a filamentous phage acquired by Yersinia pestis

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Anne eDerbise

2014-12-01

Full Text Available Yersinia pestis, the plague bacillus, has an exceptional pathogenicity for humans. The plague bacillus emerged very recently (≈3,000 years ago from the enteropathogen Y. pseudotuberculosis. Early after its emergence, Y. pestis became infected by a filamentous phage named YpfΦ. During the microevolution of the plague bacillus, the phage remained in the various lineages as an unstable extrachromosomal element. However, in the sub branch that caused the third plague pandemic, YpfΦ integrated itself into the bacterial chromosome to become a stable prophage. The genome of this phage has the same genetic organization as that of other filamentous phages such as the V. cholerae CTXΦ phage, and shares high sequence identity with the CUS-1 filamentous phage of a high-virulence E. coli K1 clone. In addition to genes involved in phage physiology, YpfΦ carries at each extremity of its genome two open reading frames with no predicted functions. This filamentous phage confers some selective properties to Y. pestis during the infectious process, which may explain why it was conserved during Y. pestis microevolution, despite its instability as an extrachromosomal element in most branches.

Twelve previously unknown phage genera are ubiquitous in global oceans.

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Holmfeldt, Karin; Solonenko, Natalie; Shah, Manesh; Corrier, Kristen; Riemann, Lasse; Verberkmoes, Nathan C; Sullivan, Matthew B

2013-07-30

Viruses are fundamental to ecosystems ranging from oceans to humans, yet our ability to study them is bottlenecked by the lack of ecologically relevant isolates, resulting in "unknowns" dominating culture-independent surveys. Here we present genomes from 31 phages infecting multiple strains of the aquatic bacterium Cellulophaga baltica (Bacteroidetes) to provide data for an underrepresented and environmentally abundant bacterial lineage. Comparative genomics delineated 12 phage groups that (i) each represent a new genus, and (ii) represent one novel and four well-known viral families. This diversity contrasts the few well-studied marine phage systems, but parallels the diversity of phages infecting human-associated bacteria. Although all 12 Cellulophaga phages represent new genera, the podoviruses and icosahedral, nontailed ssDNA phages were exceptional, with genomes up to twice as large as those previously observed for each phage type. Structural novelty was also substantial, requiring experimental phage proteomics to identify 83% of the structural proteins. The presence of uncommon nucleotide metabolism genes in four genera likely underscores the importance of scavenging nutrient-rich molecules as previously seen for phages in marine environments. Metagenomic recruitment analyses suggest that these particular Cellulophaga phages are rare and may represent a first glimpse into the phage side of the rare biosphere. However, these analyses also revealed that these phage genera are widespread, occurring in 94% of 137 investigated metagenomes. Together, this diverse and novel collection of phages identifies a small but ubiquitous fraction of unknown marine viral diversity and provides numerous environmentally relevant phage-host systems for experimental hypothesis testing.

Phages of Listeria offer novel tools for diagnostics and biocontrol

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Martin J Loessner

2014-04-01

Full Text Available Historically, bacteriophages infecting their hosts have perhaps been best known and even notorious for being a nuisance in dairy-fermentation processes. However, with the rapid progress in molecular microbiology and microbial ecology, a new dawn has risen for phages. This review will provide an overview on possible uses and applications of Listeria phages, including phage-typing, reporter phage for bacterial diagnostics, and use of phage as biocontrol agents for food safety. The use of phage-encoded enzymes such as endolysins for the detection and as antimicrobial will also be addressed. Desirable properties of candidate phages for biocontrol will be discussed. While emphasizing the enormous future potential for applications, we will also consider some of the intrinsic limitations dictated by both phage and bacterial ecology.

Engineering Circular Gliding of Actin Filaments Along Myosin-Patterned DNA Nanotube Rings To Study Long-Term Actin-Myosin Behaviors.

Science.gov (United States)

Hariadi, Rizal F; Appukutty, Abhinav J; Sivaramakrishnan, Sivaraj

2016-09-27

Nature has evolved molecular motors that are critical in cellular processes occurring over broad time scales, ranging from seconds to years. Despite the importance of the long-term behavior of molecular machines, topics such as enzymatic lifetime are underexplored due to the lack of a suitable approach for monitoring motor activity over long time periods. Here, we developed an "O"-shaped Myosin Empowered Gliding Assay (OMEGA) that utilizes engineered micron-scale DNA nanotube rings with precise arrangements of myosin VI to trap gliding actin filaments. This circular gliding assay platform allows the same individual actin filament to glide over the same myosin ensemble (50-1000 motors per ring) multiple times. First, we systematically characterized the formation of DNA nanotubes rings with 4, 6, 8, and 10 helix circumferences. Individual actin filaments glide along the nanotube rings with high processivity for up to 12.8 revolutions or 11 min in run time. We then show actin gliding speed is robust to variation in motor number and independent of ring curvature within our sample space (ring diameter of 0.5-4 μm). As a model application of OMEGA, we then analyze motor-based mechanical influence on "stop-and-go" gliding behavior of actin filaments, revealing that the stop-to-go transition probability is dependent on motor flexibility. Our circular gliding assay may provide a closed-loop platform for monitoring long-term behavior of broad classes of molecular motors and enable characterization of motor robustness and long time scale nanomechanical processes.

Efficient identification of phosphatidylserine-binding proteins by ORF phage display

International Nuclear Information System (INIS)

Caberoy, Nora B.; Zhou, Yixiong; Alvarado, Gabriela; Fan, Xianqun; Li, Wei

2009-01-01

To efficiently elucidate the biological roles of phosphatidylserine (PS), we developed open-reading-frame (ORF) phage display to identify PS-binding proteins. The procedure of phage panning was optimized with a phage clone expressing MFG-E8, a well-known PS-binding protein. Three rounds of phage panning with ORF phage display cDNA library resulted in ∼300-fold enrichment in PS-binding activity. A total of 17 PS-binding phage clones were identified. Unlike phage display with conventional cDNA libraries, all 17 PS-binding clones were ORFs encoding 13 real proteins. Sequence analysis revealed that all identified PS-specific phage clones had dimeric basic amino acid residues. GST fusion proteins were expressed for 3 PS-binding proteins and verified for their binding activity to PS liposomes, but not phosphatidylcholine liposomes. These results elucidated previously unknown PS-binding proteins and demonstrated that ORF phage display is a versatile technology capable of efficiently identifying binding proteins for non-protein molecules like PS.

The genomes and comparative genomics of Lactobacillus delbrueckii phages.

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Riipinen, Katja-Anneli; Forsman, Päivi; Alatossava, Tapani

2011-07-01

Lactobacillus delbrueckii phages are a great source of genetic diversity. Here, the genome sequences of Lb. delbrueckii phages LL-Ku, c5 and JCL1032 were analyzed in detail, and the genetic diversity of Lb. delbrueckii phages belonging to different taxonomic groups was explored. The lytic isometric group b phages LL-Ku (31,080 bp) and c5 (31,841 bp) showed a minimum nucleotide sequence identity of 90% over about three-fourths of their genomes. The genomic locations of their lysis modules were unique, and the genomes featured several putative overlapping transcription units of genes. LL-Ku and c5 virions displayed peptidoglycan hydrolytic activity associated with a ~36-kDa protein similar in size to the endolysin. Unexpectedly, the 49,433-bp genome of the prolate phage JCL1032 (temperate, group c) revealed a conserved gene order within its structural genes. Lb. delbrueckii phages representing groups a (a phage LL-H), b and c possessed only limited protein sequence homology. Genomic comparison of LL-Ku and c5 suggested that diversification of Lb. delbrueckii phages is mainly due to insertions, deletions and recombination. For the first time, the complete genome sequences of group b and c Lb. delbrueckii phages are reported.

Restoring logic and data to phage-cures for infectious disease

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Philip Serwer

2017-08-01

Full Text Available Antibiotic therapy for infectious disease is being compromised by emergence of multi-drug-resistant bacterial strains, often called superbugs. A response is to use a cocktail of several bacteria-infecting viruses (bacteriophages or phages to supplement antibiotic therapy. Use of such cocktails is called phage therapy, which has the advantage of response to bacterial resistance that is rapid and not exhaustible. A procedure of well-established success is to make cocktails from stockpiles of stored environmental phages. New phages are added to stockpiles when phage therapy becomes thwarted. The scientific subtext includes optimizing the following aspects: (1 procedure for rapidly detecting, purifying, storing and characterizing phages for optimization of phage cocktails, (2 use of directed evolution in the presence of bacteriostatic compounds to obtain phages that can be most efficiently used for therapy in the presence of these compounds, (3 phage genome sequencing technology and informatics to improve the characterization of phages, and (4 database technology to make optimal use of all relevant information and to rapidly retrieve phages for cocktails that will vary with the infection(s involved. The use of phage stockpiles has an established record, including a recent major human-therapy success by the US Navy. However, I conclude that most research is not along this track and, therefore, is not likely to lead to real world success. I find that a strong case exists for action to rectify this situation.

Synthesis of dispersive iron or iron–silver nanoparticles on engineered capsid pVIII of M13 virus with electronegative terminal peptides

Energy Technology Data Exchange (ETDEWEB)

Zhang, Shuai; Nakano, Kazuhiko; Zhang, Shu-liang; Yu, Hui-min, E-mail: yuhm@tsinghua.edu.cn [Tsinghua University, Key Laboratory of Industrial Biocatalysis of the Ministry of Education, Department of Chemical Engineering (China)

2015-10-15

M13 is a filamentous Escherichia coli virus covered with five types of capsid proteins, in which pVIII with ∼2700 copies was around the cylindered surface and pIII with five copies located at one end of the phage particle. The pIII-engineered M13 phages with enhanced binding specificity toward Fe were screened after five rounds of biopanning, and the one containing ATPTVAMSLSPL peptide at pIII-terminus was selected for mediated synthesis of zero valent (ZV) Fe nanoparticles (NPs) with the wild M13 as control. Under a reducing environment, uniformly dispersed ZVFeNPs with diameter of 5–10 nm were both synthesized and the morphologies after annealing were confirmed to be face-centered cubic type. The synthesized FeNPs mediated by the two phages showed no significant difference, revealing that the pVIII capsid did dominant contribution to metal binding in comparison with the pIII. A novel pVIII-engineered M13 containing AAEEEDPAK at terminus, named as 4ED-pVIII-M13, was constructed and it carried one more negatively charged residue than the wild one (AEGDDPAK). Metal adsorption quantification showed that the binding affinity of the 4ED-pVIII-M13 toward Ag and Ni ions improved to 62 and 18 % from original 21 and 6 %, respectively. The binding affinity toward Fe remained constant (∼85 %). ZVFe–Ag bi-NPs were successfully synthesized through mediation of 4ED-pVIII-M13. Particularly, the Fe:Ag ratio in the bi-NPs was conveniently controlled through changing the molar concentration of FeCl{sub 2} and AgNO{sub 3} solution before reduction.

Supersize me: Cronobacter sakazakii phage GAP32

Energy Technology Data Exchange (ETDEWEB)

Abbasifar, Reza; Griffiths, Mansel W. [Canadian Research Institute for Food Safety, University of Guelph, Guelph, ON, Canada N1G 2W1 (Canada); Sabour, Parviz M. [Agriculture and Agri-Food Canada, Guelph Food Research Centre, Guelph, ON, Canada N1G 5C9 (Canada); Ackermann, Hans-Wolfgang [Department of Microbiology-Infectiology and Immunology, Faculty of Medicine, Université Laval, Quebec, QC (Canada); Vandersteegen, Katrien; Lavigne, Rob [Laboratory of Gene Technology, Katholieke Universiteit Leuven, Leuven (Belgium); Noben, Jean-Paul [Biomedical Research Institute and Transnational University Limburg, School of Life Sciences, Hasselt University, Diepenbeek (Belgium); Alanis Villa, Argentina; Abbasifar, Arash [Canadian Research Institute for Food Safety, University of Guelph, Guelph, ON, Canada N1G 2W1 (Canada); Nash, John H.E. [Public Health Agency of Canada, Laboratory for Foodborne Zoonoses, Guelph, ON, Canada N1G 3W4 (Canada); Kropinski, Andrew M., E-mail: akropins@uoguelph.ca [Public Health Agency of Canada, Laboratory for Foodborne Zoonoses, Guelph, ON, Canada N1G 3W4 (Canada); Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, Canada N1G 2W1 (Canada)

2014-07-15

Cronobacter sakazakii is a Gram-negative pathogen found in milk-based formulae that causes infant meningitis. Bacteriophages have been proposed to control bacterial pathogens; however, comprehensive knowledge about a phage is required to ensure its safety before clinical application. We have characterized C. sakazakii phage vB{sub C}saM{sub G}AP32 (GAP32), which possesses the second largest sequenced phage genome (358,663 bp). A total of 571 genes including 545 protein coding sequences and 26 tRNAs were identified, thus more genes than in the smallest bacterium, Mycoplasma genitalium G37. BLASTP and HHpred searches, together with proteomic analyses reveal that only 23.9% of the putative proteins have defined functions. Some of the unique features of this phage include: a chromosome condensation protein, two copies of the large subunit terminase, a predicted signal-arrest-release lysin; and an RpoD-like protein, which is possibly involved in the switch from immediate early to delayed early transcription. Its closest relatives are all extremely large myoviruses, namely coliphage PBECO4 and Klebsiella phage vB{sub K}leM-RaK2, with whom it shares approximately 44% homologous proteins. Since the homologs are not evenly distributed, we propose that these three phages belong to a new subfamily. - Highlights: • Cronobacter sakazakii phage vB{sub C}saM{sub G}AP32 has a genome of 358,663 bp. • It encodes 545 proteins which is more than Mycoplasma genitalium G37. • It is a member of the Myoviridae. • It is peripherally related to coliphage PBECO4 and Klebsiella phage vB{sub K}leM-RaK2. • GAP32 encodes a chromosome condensation protein.

Coevolution of CRISPR bacteria and phage in 2 dimensions

Science.gov (United States)

Han, Pu; Deem, Michael

2014-03-01

CRISPR (cluster regularly interspaced short palindromic repeats) is a newly discovered adaptive, heritable immune system of prokaryotes. It can prevent infection of prokaryotes by phage. Most bacteria and almost all archae have CRISPR. The CRISPR system incorporates short nucleotide sequences from viruses. These incorporated sequences provide a historical record of the host and predator coevolution. We simulate the coevolution of bacteria and phage in 2 dimensions. Each phage has multiple proto-spacers that the bacteria can incorporate. Each bacterium can store multiple spacers in its CRISPR. Phages can escape recognition by the CRISPR system via point mutation or recombination. We will discuss the different evolutionary consequences of point mutation or recombination on the coevolution of bacteria and phage. We will also discuss an intriguing ``dynamic phase transition'' in the number of phage as a function of time and mutation rate. We will show that due to the arm race between phages and bacteria, the frequency of spacers and proto-spacers in a population can oscillate quite rapidly.

Influence of environmental factors on phage-bacteria interaction and on the efficacy and infectivity of phage P100

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Susanne Fister

2016-07-01

Full Text Available When using bacteriophages to control food-borne bacteria in food production plants and processed food, it is crucial to consider that environmental conditions influence their stability. These conditions can also affect the physiological state of bacteria and consequently host-virus interaction and the effectiveness of the phage ability to reduce bacteria numbers. In this study we investigated the stability, binding and replication capability of phage P100 and its efficacy to control L. monocytogenes under conditions typically encountered in dairy plants. The influences of SDS, Lutensol AO 7, salt, smear water and different temperatures were investigated. Results indicate that phage P100 is stable and able to bind to the host under most conditions tested. Replication was dependent upon the growth of L. monocytogenes and efficacy was higher when bacterial growth was reduced by certain environmental conditions. In long-term experiments at different temperatures phages were initially able to reduce bacteria up to seven log10 units after two weeks at 4 °C. However, thereafter re-growth and development of phage-resistant L. monocytogenes isolates were encountered.

Phage therapy reduces Campylobacter jejuni colonization in broilers

NARCIS (Netherlands)

Wagenaar, J.A.; Bergen, van M.A.P.; Mueller, M.A.; Wassenaar, T.M.; Carlton, R.M.

2005-01-01

The effect of phage therapy in the control of Campylobacter jejuni colonization in young broilers, either as a preventive or a therapeutic measure, was tested. A prevention group was infected with C. jejuni at day 4 of a 10-day phage treatment. A therapeutic group was phage treated for 6 days,

Molecular Characterization of Three Lactobacillus delbrueckii subsp. bulgaricus Phages

Science.gov (United States)

Casey, Eoghan; Mahony, Jennifer; O'Connell-Motherway, Mary; Bottacini, Francesca; Cornelissen, Anneleen; Neve, Horst; Heller, Knut J.; Noben, Jean-Paul; Dal Bello, Fabio

2014-01-01

In this study, three phages infecting Lactobacillus delbrueckii subsp. bulgaricus, named Ld3, Ld17, and Ld25A, were isolated from whey samples obtained from various industrial fermentations. These phages were further characterized in a multifaceted approach: (i) biological and physical characterization through host range analysis and electron microscopy; (ii) genetic assessment through genome analysis; (iii) mass spectrometry analysis of the structural components of the phages; and (iv), for one phage, transcriptional analysis by Northern hybridization, reverse transcription-PCR, and primer extension. The three obtained phage genomes display high levels of sequence identity to each other and to genomes of the so-called group b L. delbrueckii phages c5, LL-Ku, and phiLdb, where some of the observed differences are believed to be responsible for host range variations. PMID:25002431

Scaling Up: Adapting a Phage-Hunting Course to Increase Participation of First-Year Students in Research.

Science.gov (United States)

Staub, Nancy L; Poxleitner, Marianne; Braley, Amanda; Smith-Flores, Helen; Pribbenow, Christine M; Jaworski, Leslie; Lopatto, David; Anders, Kirk R

2016-01-01

Authentic research experiences are valuable components of effective undergraduate education. Research experiences during the first years of college are especially critical to increase persistence in science, technology, engineering, and mathematics fields. The Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) model provides a high-impact research experience to first-year students but is usually available to a limited number of students, and its implementation is costly in faculty time and laboratory space. To offer a research experience to all students taking introductory biology at Gonzaga University (n = 350/yr), we modified the traditional two-semester SEA-PHAGES course by streamlining the first-semester Phage Discovery lab and integrating the second SEA-PHAGES semester into other courses in the biology curriculum. Because most students in the introductory course are not biology majors, the Phage Discovery semester may be their only encounter with research. To discover whether students benefit from the first semester alone, we assessed the effects of the one-semester Phage Discovery course on students' understanding of course content. Specifically, students showed improvement in knowledge of bacteriophages, lab math skills, and understanding experimental design and interpretation. They also reported learning gains and benefits comparable with other course-based research experiences. Responses to open-ended questions suggest that students experienced this course as a true undergraduate research experience. © 2016 N. L. Staub et al. CBE—Life Sciences Education © 2016 The American Society for Cell Biology. This article is distributed by The American Society for Cell Biology under license from the author(s). It is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Characterisation of a novel enterobacteria phage, CAjan, isolated from rat faeces.

Science.gov (United States)

Carstens, Alexander B; Kot, Witold; Lametsch, Rene; Neve, Horst; Hansen, Lars H

2016-08-01

In this study, we describe the isolation and characterisation of the novel enterobacteria phage CAjan. This phage belongs to the order Caudovirales and the family Siphoviridae. The phage possesses a linear, double-stranded DNA genome consisting of 59,670 bp with a G+C content of 44.7 % and 91 predicted open reading frames (ORFs). Putative functions were assigned to 39 of the ORFs (37.4 %). The phage structural genes were furthermore functionally characterised by LC MS/MS. CAjan, together with Escherichia phage Seurat and Escherichia phage slur01, represent a novel and genetically distinct clade of Siphoviridae phages that could be considered to constitute a new phage genus. Despite limited sequence similarity, the phages in this group share a number of other common features, including genome structure and the presence of queuosine biosynthesis genes.

Advanced Analysis to Distinguish between Physical Decrease and Inactivation of Viable Phages in Aerosol by Quantitating Phage-Specific Particles.

Science.gov (United States)

Shimasaki, Noriko; Nojima, Yasuhiro; Sakakibara, Masaya; Kikuno, Ritsuko; Iizuka, Chiori; Okaue, Akira; Okuda, Shunji; Shinohara, Katsuaki

2018-01-01

　Recent studies have investigated the efficacy of air-cleaning products against pathogens in the air. A standard method to evaluate the reduction in airborne viruses caused by an air cleaner has been established using a safe bacteriophage instead of pathogenic viruses; the reduction in airborne viruses is determined by counting the number of viable airborne phages by culture, after operating the air cleaner. The reduction in the number of viable airborne phages could be because of "physical decrease" or "inactivation". Therefore, to understand the mechanism of reduction correctly, an analysis is required to distinguish between physical decrease and inactivation. The purpose of this study was to design an analysis to distinguish between the physical decrease and inactivation of viable phi-X174 phages in aerosols. We established a suitable polymerase chain reaction (PCR) system by selecting an appropriate primer-probe set for PCR and validating the sensitivity, linearity, and specificity of the primer-probe set to robustly quantify phi-X174-specific airborne particles. Using this quantitative PCR system and culture assay, we performed a behavior analysis of the phage aerosol in a small chamber (1 m 3 ) at different levels of humidity, as humidity is known to affect the number of viable airborne phages. The results revealed that the reduction in the number of viable airborne phages was caused not only by physical decrease but also by inactivation under particular levels of humidity. Our study could provide an advanced analysis to differentiate between the physical decrease and inactivation of viable airborne phages.

Therapeutic use of chimeric bacteriophage (phage) lysins in staphylococcal endophthalmitis

Science.gov (United States)

Purpose: Phage endolysins are peptidoglycan hydrolases that are produced at the end of the phage lytic cycle to digest the host bacterial cell wall, facilitating the release of mature phage progeny. The aim of this study is to determine the antimicrobial activity of chimeric phage lysins against cli...

Molecular characterization of three Lactobacillus delbrueckii subsp. bulgaricus phages.

Science.gov (United States)

Casey, Eoghan; Mahony, Jennifer; O'Connell-Motherway, Mary; Bottacini, Francesca; Cornelissen, Anneleen; Neve, Horst; Heller, Knut J; Noben, Jean-Paul; Dal Bello, Fabio; van Sinderen, Douwe

2014-09-01

In this study, three phages infecting Lactobacillus delbrueckii subsp. bulgaricus, named Ld3, Ld17, and Ld25A, were isolated from whey samples obtained from various industrial fermentations. These phages were further characterized in a multifaceted approach: (i) biological and physical characterization through host range analysis and electron microscopy; (ii) genetic assessment through genome analysis; (iii) mass spectrometry analysis of the structural components of the phages; and (iv), for one phage, transcriptional analysis by Northern hybridization, reverse transcription-PCR, and primer extension. The three obtained phage genomes display high levels of sequence identity to each other and to genomes of the so-called group b L. delbrueckii phages c5, LL-Ku, and phiLdb, where some of the observed differences are believed to be responsible for host range variations. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

In Vivo Imaging of Molecularly Targeted Phage

Directory of Open Access Journals (Sweden)

Kimberly A. Kelly

2006-12-01

Full Text Available Rapid identification of in vivo affinity ligands would have far-reaching applications for imaging specific molecular targets, in vivo systems imaging, and medical use. We have developed a high-throughput method for identifying and optimizing ligands to map and image biologic targets of interest in vivo. We directly labeled viable phage clones with far-red fluorochromes and comparatively imaged them in vivo by multichannel fluorescence ratio imaging. Using Secreted Protein Acidic and Rich in Cysteine (osteonectin and vascular cell adhesion molecule-1 as model targets, we show that: 1 fluorescently labeled phage retains target specificity on labeling; 2 in vivo distribution can be quantitated (detection thresholds of ~ 300 phage/mm3 tissue throughout the entire depth of the tumor using fluorescent tomographic imaging; and 3 fluorescently labeled phage itself can serve as a replenishable molecular imaging agent. The described method should find widespread application in the rapid in vivo discovery and validation of affinity ligands and, importantly, in the use of fluorochrome-labeled phage clones as in vivo imaging agents.

How to Name and Classify Your Phage: An Informal Guide

Directory of Open Access Journals (Sweden)

Evelien Adriaenssens

2017-04-01

Full Text Available With this informal guide, we try to assist both new and experienced phage researchers through two important stages that follow phage discovery; that is, naming and classification. Providing an appropriate name for a bacteriophage is not as trivial as it sounds, and the effects might be long-lasting in databases and in official taxon names. Phage classification is the responsibility of the Bacterial and Archaeal Viruses Subcommittee (BAVS of the International Committee on the Taxonomy of Viruses (ICTV. While the BAVS aims at providing a holistic approach to phage taxonomy, for individual researchers who have isolated and sequenced a new phage, this can be a little overwhelming. We are now providing these researchers with an informal guide to phage naming and classification, taking a “bottom-up” approach from the phage isolate level.

Engineering bacteriophage for a pragmatic low-resource setting bacterial diagnostic platform.

Science.gov (United States)

Talbert, Joey N; Alcaine, Samuel D; Nugen, Sam R

2016-04-01

Bacteriophages represent multifaceted building blocks that can be incorporated as substitutes for, or in unison with other detection methods, to create powerful new diagnostics for the detection of bacteria. The ease of phage manipulation, production, and detection speed clearly highlights that there remains unrealized opportunities to leverage these phage-based components in diagnostics amenable to resource-limited settings. The passage of regulations like the Food Safety Modernization act, and the ever increasing extent of global trade and travel, will create further demand for these types of diagnostics. While phage-based diagnostics have begun to entering the market place, further research is needed to ensure the potential benefits of phage-based technologies for public health are fully realized. We are just beginning to explore the possibilities that phage-based detection can offer us in the future. The combination of engineered phages as well as engineered enzymes could result in ultrasensitive detection systems for low-resource settings. Because the reporter enzyme is synthesized in vivo, we need to consider the options outside of normal enzyme reporters. In this case, common enzyme issues such as purification and long-term stability are less important. Phage-based diagnostics were conceptualized from out-of-the box thinking and the evolution of these systems should be as well.

Lysis-deficient phages as novel therapeutic agents for controlling bacterial infection

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Kempashanaiah Nanjundappa

2011-08-01

Full Text Available Abstract Background Interest in phage therapy has grown over the past decade due to the rapid emergence of antibiotic resistance in bacterial pathogens. However, the use of bacteriophages for therapeutic purposes has raised concerns over the potential for immune response, rapid toxin release by the lytic action of phages, and difficulty in dose determination in clinical situations. A phage that kills the target cell but is incapable of host cell lysis would alleviate these concerns without compromising efficacy. Results We developed a recombinant lysis-deficient Staphylococcus aureus phage P954, in which the endolysin gene was rendered nonfunctional by insertional inactivation. P954, a temperate phage, was lysogenized in S. aureus strain RN4220. The native endolysin gene on the prophage was replaced with an endolysin gene disrupted by the chloramphenicol acetyl transferase (cat gene through homologous recombination using a plasmid construct. Lysogens carrying the recombinant phage were detected by growth in presence of chloramphenicol. Induction of the recombinant prophage did not result in host cell lysis, and the phage progeny were released by cell lysis with glass beads. The recombinant phage retained the endolysin-deficient genotype and formed plaques only when endolysin was supplemented. The host range of the recombinant phage was the same as that of the parent phage. To test the in vivo efficacy of the recombinant endolysin-deficient phage, immunocompromised mice were challenged with pathogenic S. aureus at a dose that results in 80% mortality (LD80. Treatment with the endolysin-deficient phage rescued mice from the fatal S. aureus infection. Conclusions A recombinant endolysin-deficient staphylococcal phage has been developed that is lethal to methicillin-resistant S. aureus without causing bacterial cell lysis. The phage was able to multiply in lytic mode utilizing a heterologous endolysin expressed from a plasmid in the propagation host

Mutagenesis of lambda phage by tif-expression or host-irradiation functions is largely independent of damage in the phage DNA

International Nuclear Information System (INIS)

Von Wright, A.; Bridges, B.A.

1980-01-01

The survival and mutagenesis of UV-irradiated phage lambda, as well as bacterial mutagenesis, are enhanced in tif mutants of Escherichia coli when these strains are grown at 43 0 C (Castellazzi et al., 1972). This was interpreted on the basis of a hypothesis (the SOS hypothesis) according to which the UV-inducible phenomena connected with reactivation and mutagenesis of UV-irradiated bacteriophages (Weigle, 1953; Radman, 1975) are constitutively expressed in tif-bacteria at high temperature (Witkin, 1974). In unpublished experiments with phage T3 we found that the survival of UV-irradiated phage is also better at 43 0 C than at 32 0 C in tif + cells and this made us reexamine the significance and nature of tif expression and examine its effects on both unirradiated and UV-irradiated phage lambda. Our results indicate that tif-induced mutagenesis and possibly reactivation of UV-irradiated phage lambda should be reinterpreted. (orig./AJ)

The phage-host arms race: Shaping the evolution of microbes

Energy Technology Data Exchange (ETDEWEB)

Stern, Adi [Weizmann Inst. of Science, Rehovot (Israel). Dept. of Molecular Genetics; Sorek, Rotem [Weizmann Inst. of Science, Rehovot (Israel). Dept. of Molecular Genetics

2010-10-26

Bacteria, the most abundant organisms on the planet, are outnumbered by a factor of 10 to 1 by phages that infect them. Faced with the rapid evolution and turnover of phage particles, bacteria have evolved various mechanisms to evade phage infection and killing, leading to an evolutionary arms race. The extensive co-evolution of both phage and host has resulted in considerable diversity on the part of both bacterial and phage defensive and offensive strategies. In this paper, we discuss the unique and common features of phage resistance mechanisms and their role in global biodiversity. Finally, the commonalities between defense mechanisms suggest avenues for the discovery of novel forms of these mechanisms based on their evolutionary traits.

Sensitivity of RF-driven Plasma Filaments to Trace Gases

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Burin, M. J.; Czarnocki, C. J.; Czarnocki, K.; Zweben, S. J.; Zwicker, A.

2011-10-01

Filamentary structures have been observed in many types of plasma discharges in both natural (e.g. lightning) and industrial systems (e.g. dielectric barrier discharges). Recent progress has been made in characterizing these structures, though various aspects of their essential physics remain unclear. A common example of this phenomenon can be found within a toy plasma globe (or plasma ball), wherein a primarily neon gas mixture near atmospheric pressure clearly and aesthetically displays filamentation. Recent work has provided the first characterization of these plasma globe filaments [Campanell et al., Physics of Plasmas 2010], where it was noticed that discharges of pure gases tend not to produce filaments. We have extended this initial work to investigate in greater detail the dependence of trace gases on filamentation within a primarily Neon discharge. Our preliminary results using a custom globe apparatus will be presented, along with some discussion of voltage dependencies. Newly supported by the NSF/DOE Partnership in Basic Plasma Science and Engineering.

Bacterial filamentation accelerates colonization of adhesive spots embedded in biopassive surfaces

International Nuclear Information System (INIS)

Möller, Jens; Emge, Philippe; Vizcarra, Ima Avalos; Kollmannsberger, Philip; Vogel, Viola

2013-01-01

Sessile bacteria adhere to engineered surfaces and host tissues and pose a substantial clinical and economical risk when growing into biofilms. Most engineered and biological interfaces are of chemically heterogeneous nature and provide adhesive islands for bacterial attachment and growth. To mimic either defects in a surface coating of biomedical implants or heterogeneities within mucosal layers (Peyer's patches), we embedded micrometre-sized adhesive islands in a poly(ethylene glycol) biopassive background. We show experimentally and computationally that filamentation of Escherichia coli can significantly accelerate the bacterial surface colonization under physiological flow conditions. Filamentation can thus provide an advantage to a bacterial population to bridge non-adhesive distances exceeding 5 μm. Bacterial filamentation, caused by blocking of bacterial division, is common among bacterial species and can be triggered by environmental conditions or antibiotic treatment. While great awareness exists that the build-up of antibiotic resistance serves as intrinsic survival strategy, we show here that antibiotic treatment can actually promote surface colonization by triggering filamentation, which in turn prevents daughter cells from being washed away. Our combined microfabrication and computational approaches provide quantitative insights into mechanisms that enable biofouling of biopassive surfaces with embedded adhesive spots, even for spot distances that are multiples of the bacterial length. (paper)

Bacterial filamentation accelerates colonization of adhesive spots embedded in biopassive surfaces

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Möller, Jens; Emge, Philippe; Avalos Vizcarra, Ima; Kollmannsberger, Philip; Vogel, Viola

2013-12-01

Sessile bacteria adhere to engineered surfaces and host tissues and pose a substantial clinical and economical risk when growing into biofilms. Most engineered and biological interfaces are of chemically heterogeneous nature and provide adhesive islands for bacterial attachment and growth. To mimic either defects in a surface coating of biomedical implants or heterogeneities within mucosal layers (Peyer's patches), we embedded micrometre-sized adhesive islands in a poly(ethylene glycol) biopassive background. We show experimentally and computationally that filamentation of Escherichia coli can significantly accelerate the bacterial surface colonization under physiological flow conditions. Filamentation can thus provide an advantage to a bacterial population to bridge non-adhesive distances exceeding 5 μm. Bacterial filamentation, caused by blocking of bacterial division, is common among bacterial species and can be triggered by environmental conditions or antibiotic treatment. While great awareness exists that the build-up of antibiotic resistance serves as intrinsic survival strategy, we show here that antibiotic treatment can actually promote surface colonization by triggering filamentation, which in turn prevents daughter cells from being washed away. Our combined microfabrication and computational approaches provide quantitative insights into mechanisms that enable biofouling of biopassive surfaces with embedded adhesive spots, even for spot distances that are multiples of the bacterial length.

Marine phages as excellent tracers for reactive colloidal transport in porous media

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Ghanem, Nawras; Chatzinotas, Antonis; Harms, Hauke; Wick, Lukas Y.

2016-04-01

Question: Here we evaluate marine phages as specific markers of hydrological flow and reactive transport of colloidal particles in the Earth's critical zone (CZ). Marine phages and their bacterial hosts are naturally absent in the CZ, and can be detected with extremely high sensitivity. In the framework of the DFG Collaborative Research Center AquaDiva, we asked the following questions: (1) Are marine phages useful specific markers of hydrological flow and reactive transport in porous media? and (2) Which phage properties are relevant drivers for the transport of marine phages in porous media? Methods: Seven marine phages from different families (as well two commonly used terrestrial phages) were selected based on their morphology, size and physico-chemical surface properties (surface charge and hydrophobicity). Phage properties were assessed by electron microscopy, dynamic light scattering and water contact angle analysis (CA). Sand-filled laboratory percolation columns were used to study transport. The breakthrough curves of the phages were analyzed using the clean bed filtration theory and the XDLVO theory of colloid stability, respectively. Phages were quantified by a modified high- throughput plaque assay and a culture-independent particle counting method approach. Results: Our data show that most marine tested phages exhibited highly variable transport rates and deposition efficiency, yet generally high colloidal stability and viability. We find that size, morphology and hydrophobicity are key factors shaping the transport efficiency of phages. Differing deposition efficiencies of the phages were also supported by calculated XDLVO interaction energy profile. Conclusion: Marine phages have a high potential for the use as sensitive tracers in terrestrial habitats with their surface properties playing a crucial role for their transport. Marine phages however, exhibit differences in their deposition efficiency depending on their morphology, hydrophobicity and

An improved plating assay for determination of phage titer

African Journals Online (AJOL)

RACHEL

antibiotics to control bacterial infections in swine (Thacker,. 2014). Phage therapy is re-valued by researchers to combat the growing menace of antibiotic-resistant infections (Torres-Barceló and Hochberg, 2016). Determination of phage titer in a sample is a key step in the study of the phage involved. It is very important to.

Phage lytic enzymes: a history.

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Trudil, David

2015-02-01

There are many recent studies regarding the efficacy of bacteriophage-related lytic enzymes: the enzymes of 'bacteria-eaters' or viruses that infect bacteria. By degrading the cell wall of the targeted bacteria, these lytic enzymes have been shown to efficiently lyse Gram-positive bacteria without affecting normal flora and non-related bacteria. Recent studies have suggested approaches for lysing Gram-negative bacteria as well (Briersa Y, et al., 2014). These enzymes include: phage-lysozyme, endolysin, lysozyme, lysin, phage lysin, phage lytic enzymes, phageassociated enzymes, enzybiotics, muralysin, muramidase, virolysin and designations such as Ply, PAE and others. Bacteriophages are viruses that kill bacteria, do not contribute to antimicrobial resistance, are easy to develop, inexpensive to manufacture and safe for humans, animals and the environment. The current focus on lytic enzymes has been on their use as anti-infectives in humans and more recently in agricultural research models. The initial translational application of lytic enzymes, however, was not associated with treating or preventing a specific disease but rather as an extraction method to be incorporated in a rapid bacterial detection assay (Bernstein D, 1997).The current review traces the translational history of phage lytic enzymes-from their initial discovery in 1986 for the rapid detection of group A streptococcus in clinical specimens to evolving applications in the detection and prevention of disease in humans and in agriculture.

Ultraviolet inactivation and photoreactivation of the cholera phage 'Kappa'

International Nuclear Information System (INIS)

Samad, S.A.; Bhattacharyya, S.C.; Chatterjee, S.N.

1987-01-01

The lysogenic cholera phage, 'Kappa' is some ten to twenty folds more resistant to UV (254 nm) than are most of the T. phages of E. coli, or the cholera phage PL 163/10, or the host V. cholerae strain H218 Sm r , the 37% (D 37 ) and 10% (D 10 ) survival doses being 255.8 J/m 2 and 633.6 J/m 2 respectively. The UV-irradiated 'Kappa' phages could be photoreactivated in the host V. cholerae strain H218 Sm r to a maximum extent of 40%. The removal of the number of lethal hits per phage by the survival-enhancement treatment (photoreactivation) with time followed an exponential relation, the constant probability of removal of lethal hit per unit time being 2.8x10 -2 min -1 . The UV-irradiated phages could also be Weigle reactivated in the host strain of H218 Sm r by a small degree, the maximum reactivation factor (ratio of survivals in UV-irradiated and non-irradiated hosts) being 1.50. (orig.)

Staphylococcus aureus phage types and their correlation to antibiotic resistance

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Mehndiratta P

2010-10-01

Full Text Available Context: Staphylococcus aureus is one of the most devastating human pathogen. The organism has a differential ability to spread and cause outbreak of infections. Characterization of these strains is important to control the spread of infection in the hospitals as well as in the community. Aim: To identify the currently existing phage groups of Staphylococcus aureus, their prevalence and resistance to antibiotics. Materials and Methods: Study was undertaken on 252 Staphylococcus aureus strains isolated from clinical samples. Strains were phage typed and their resistance to antibiotics was determined following standard microbiological procedures. Statistical Analysis: Chi square test was used to compare the antibiotic susceptibility between methicillin resistant Staph. aureus (MRSA and methicillin sensitive S. aureus (MSSA strains. Results: Prevalence of MRSA and MSSA strains was found to be 29.36% and 70.65% respectively. Of these 17.56% of MRSA and 40.44% of MSSA strains were community acquired. All the MSSA strains belonging to phage type 81 from the community were sensitive to all the antibiotics tested including clindamycin and were resistant to penicillin. Forty five percent strains of phage group III and 39% of non-typable MRSA strains from the hospital were resistant to multiple antibiotics. Conclusion: The study revealed that predominant phage group amongst MRSA strains was phage group III and amongst MSSA from the community was phage group NA (phage type 81. MSSA strains isolated from the community differed significantly from hospital strains in their phage type and antibiotic susceptibility. A good correlation was observed between community acquired strains of phage type 81 and sensitivity to gentamycin and clindamycin.

Non-targeted mutagenesis of unirradiated lambda phage in Escherichia coli

International Nuclear Information System (INIS)

Wood, R.D.; Hutchinson, F.

1984-01-01

Non-targeted mutagenesis of lambda phage by ultraviolet light is the increase over background mutagenesis when non-irradiated phage are grown in irradiated Escherichia coli host cells. Such mutagenesis is caused by different processes from targeted mutagenesis, in which mutations in irradiated phage are correlated with photoproducts in the phage DNA. Non-irradiated phage grown in heavily irradiated uvr + host cells showed non-targeted mutations, which were 3/4 frameshifts, whereas targeted mutations were 2/3 transitions. For non-targeted mutagenesis in heavily irradiated host cells, there were one or two mutant phage per mutant burst. From the results of a series of experiments with various mutant host cells, a major pathway of non-targeted mutagenesis by ultraviolet light was proposed which acts in addition to ''SOS induction''. This pathway involves binding of the enzyme DNA polymerase I to damaged genomic DNA, and low polymerase activity leads to frameshift mutations during semiconservative DNA replication. The data suggest that this process will play a much smaller role in ultraviolet mutagenesis of the bacterial genome than it does in the mutagenesis of lambda phage. (author)

Mutation-specific effects on thin filament length in thin filament myopathy.

Science.gov (United States)

Winter, Josine M de; Joureau, Barbara; Lee, Eun-Jeong; Kiss, Balázs; Yuen, Michaela; Gupta, Vandana A; Pappas, Christopher T; Gregorio, Carol C; Stienen, Ger J M; Edvardson, Simon; Wallgren-Pettersson, Carina; Lehtokari, Vilma-Lotta; Pelin, Katarina; Malfatti, Edoardo; Romero, Norma B; Engelen, Baziel G van; Voermans, Nicol C; Donkervoort, Sandra; Bönnemann, C G; Clarke, Nigel F; Beggs, Alan H; Granzier, Henk; Ottenheijm, Coen A C

2016-06-01

Thin filament myopathies are among the most common nondystrophic congenital muscular disorders, and are caused by mutations in genes encoding proteins that are associated with the skeletal muscle thin filament. Mechanisms underlying muscle weakness are poorly understood, but might involve the length of the thin filament, an important determinant of force generation. We investigated the sarcomere length-dependence of force, a functional assay that provides insights into the contractile strength of muscle fibers as well as the length of the thin filaments, in muscle fibers from 51 patients with thin filament myopathy caused by mutations in NEB, ACTA1, TPM2, TPM3, TNNT1, KBTBD13, KLHL40, and KLHL41. Lower force generation was observed in muscle fibers from patients of all genotypes. In a subset of patients who harbor mutations in NEB and ACTA1, the lower force was associated with downward shifted force-sarcomere length relations, indicative of shorter thin filaments. Confocal microscopy confirmed shorter thin filaments in muscle fibers of these patients. A conditional Neb knockout mouse model, which recapitulates thin filament myopathy, revealed a compensatory mechanism; the lower force generation that was associated with shorter thin filaments was compensated for by increasing the number of sarcomeres in series. This allowed muscle fibers to operate at a shorter sarcomere length and maintain optimal thin-thick filament overlap. These findings might provide a novel direction for the development of therapeutic strategies for thin filament myopathy patients with shortened thin filament lengths. Ann Neurol 2016;79:959-969. © 2016 American Neurological Association.

Analysis of high-throughput sequencing and annotation strategies for phage genomes.

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Matthew R Henn

Full Text Available BACKGROUND: Bacterial viruses (phages play a critical role in shaping microbial populations as they influence both host mortality and horizontal gene transfer. As such, they have a significant impact on local and global ecosystem function and human health. Despite their importance, little is known about the genomic diversity harbored in phages, as methods to capture complete phage genomes have been hampered by the lack of knowledge about the target genomes, and difficulties in generating sufficient quantities of genomic DNA for sequencing. Of the approximately 550 phage genomes currently available in the public domain, fewer than 5% are marine phage. METHODOLOGY/PRINCIPAL FINDINGS: To advance the study of phage biology through comparative genomic approaches we used marine cyanophage as a model system. We compared DNA preparation methodologies (DNA extraction directly from either phage lysates or CsCl purified phage particles, and sequencing strategies that utilize either Sanger sequencing of a linker amplification shotgun library (LASL or of a whole genome shotgun library (WGSL, or 454 pyrosequencing methods. We demonstrate that genomic DNA sample preparation directly from a phage lysate, combined with 454 pyrosequencing, is best suited for phage genome sequencing at scale, as this method is capable of capturing complete continuous genomes with high accuracy. In addition, we describe an automated annotation informatics pipeline that delivers high-quality annotation and yields few false positives and negatives in ORF calling. CONCLUSIONS/SIGNIFICANCE: These DNA preparation, sequencing and annotation strategies enable a high-throughput approach to the burgeoning field of phage genomics.

Spatial separation of photosynthesis and ethanol production by cell type-specific metabolic engineering of filamentous cyanobacteria.

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Ehira, Shigeki; Takeuchi, Takuto; Higo, Akiyoshi

2018-02-01

Cyanobacteria, which perform oxygenic photosynthesis, have drawn attention as hosts for the direct production of biofuels and commodity chemicals from CO 2 and H 2 O using light energy. Although cyanobacteria capable of producing diverse chemicals have been generated by metabolic engineering, anaerobic non-photosynthetic culture conditions are often necessary for their production. In this study, we conducted cell type-specific metabolic engineering of the filamentous cyanobacterium Anabaena sp. PCC 7120, which forms a terminally differentiated cell called a heterocyst with a semi-regular spacing of 10-15 cells. Because heterocysts are specialized cells for nitrogen fixation, the intracellular oxygen level of heterocysts is maintained very low even when adjacent cells perform oxygenic photosynthesis. Pyruvate decarboxylase of Zymomonas mobilis and alcohol dehydrogenase of Synechocystis sp. PCC 6803 were exclusively expressed in heterocysts. Ethanol production was concomitant with nitrogen fixation in genetically engineered Anabaena sp. PCC 7120. Engineering of carbon metabolism in heterocysts improved ethanol production, and strain ET14, with an extra copy of the invB gene expressed from a heterocyst-specific promoter, produced 130.9 mg L -1 of ethanol after 9 days. ET14 produced 1681.9 mg L -1 of ethanol by increasing the CO 2 supply. Ethanol production per heterocyst cell was approximately threefold higher than that per cell of unicellular cyanobacterium. This study demonstrates the potential of heterocysts for anaerobic production of biofuels and commodity chemicals under oxygenic photosynthetic conditions.

Complete genome sequences of three Campylobacter jejuni phage-propagating strains

Science.gov (United States)

Bacteriophage therapy has the potential to reduce Campylobacter jejuni numbers in livestock, but requires a detailed understanding of phage-host interactions. Some C. jejuni strains are readily infected by certain phages, and are thus designated as phage-propagating strains. Here we report the compl...

Non-targeted mutagenesis of unirradiated lambda phage in Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Wood, R.D.; Hutchinson, F. (Yale Univ., New Haven, CT (USA). Dept. of Molecular Biophysics and Biochemistry)

1984-03-05

Non-targeted mutagenesis of lambda phage by ultraviolet light is the increase over background mutagenesis when non-irradiated phage are grown in irradiated Escherichia coli host cells. Such mutagenesis is caused by different processes from targeted mutagenesis, in which mutations in irradiated phage are correlated with photoproducts in the phage DNA. Non-irradiated phage grown in heavily irradiated uvr/sup +/ host cells showed non-targeted mutations, which were 3/4 frameshifts, whereas targeted mutations were 2/3 transitions. For non-targeted mutagenesis in heavily irradiated host cells, there were one or two mutant phage per mutant burst. From the results of a series of experiments with various mutant host cells, a major pathway of non-targeted mutagenesis by ultraviolet light was proposed which acts in addition to ''SOS induction''. This pathway involves binding of the enzyme DNA polymerase I to damaged genomic DNA, and low polymerase activity leads to frameshift mutations during semiconservative DNA replication. The data suggest that this process will play a much smaller role in ultraviolet mutagenesis of the bacterial genome than it does in the mutagenesis of lambda phage.

Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis.

Science.gov (United States)

Furusawa, Takaaki; Iwano, Hidetomo; Hiyashimizu, Yutaro; Matsubara, Kazuki; Higuchi, Hidetoshi; Nagahata, Hajime; Niwa, Hidekazu; Katayama, Yoshinari; Kinoshita, Yuta; Hagiwara, Katsuro; Iwasaki, Tomohito; Tanji, Yasunori; Yokota, Hiroshi; Tamura, Yutaka

2016-09-01

Bacterial keratitis of the horse is mainly caused by staphylococci, streptococci, and pseudomonads. Of these bacteria, Pseudomonas aeruginosa sometimes causes rapid corneal corruption and, in some cases, blindness. Antimicrobial resistance can make treatment very difficult. Therefore, new strategies to control bacterial infection are required. A bacteriophage (phage) is a virus that specifically infects and kills bacteria. Since phage often can lyse antibiotic-resistant bacteria because the killing mechanism is different, we examined the use of phage to treat horse bacterial keratitis. We isolated Myoviridae or Podoviridae phages, which together have a broad host range. They adsorb efficiently to host bacteria; more than 80% of the ΦR18 phage were adsorbed to host cells after 30 s. In our keratitis mouse model, the administration of phage within 3 h also could kill bacteria and suppress keratitis. A phage multiplicity of infection of 100 times the host bacterial number could kill host bacteria effectively. A cocktail of two phages suppressed bacteria in the keratitis model mouse. These data demonstrated that the phages in this study could completely prevent the keratitis caused by P. aeruginosa in a keratitis mouse model. Furthermore, these results suggest that phage may be a more effective prophylaxis for horse keratitis than the current preventive use of antibiotics. Such treatment may reduce the use of antibiotics and therefore antibiotic resistance. Further studies are required to assess phage therapy as a candidate for treatment of horse keratitis. Antibiotic-resistant bacteria are emerging all over the world. Bacteriophages have great potential for resolution of this problem. A bacteriophage, or phage, is a virus that infects bacteria specifically. As a novel therapeutic strategy against racehorse keratitis caused by Pseudomonas aeruginosa, we propose the application of phages for treatment. Phages isolated in this work had in vitro effectiveness for a broad

Evolution of phage display technology: from discovery to application.

Science.gov (United States)

Rahbarnia, Leila; Farajnia, Safar; Babaei, Hossein; Majidi, Jafar; Veisi, Kamal; Ahmadzadeh, Vahideh; Akbari, Bahman

2017-03-01

Phage display technology as a selection-based system is an attractive method for evolution of new biological drugs. Unique ability of phage libraries for displaying proteins on bacteriophage surfaces enable them to make a major contribution in diverse fields of researches related to the diagnosis and therapy of diseases. One of the great challenges facing researchers is the modification of phage display technology and the development of new applications. This article reviews the molecular basis of phage display library, and summarizes the novel and specific applications of this technique in the field of biological drugs development including therapeutic antibodies, peptides, vaccines, and catalytic antibodies.

Complete Genome Sequence of Pseudomonas aeruginosa Phage AAT-1.

Science.gov (United States)

Andrade-Domínguez, Andrés; Kolter, Roberto

2016-08-25

Aspects of the interaction between phages and animals are of interest and importance for medical applications. Here, we report the genome sequence of the lytic Pseudomonas phage AAT-1, isolated from mammalian serum. AAT-1 is a double-stranded DNA phage, with a genome of 57,599 bp, containing 76 predicted open reading frames. Copyright © 2016 Andrade-Domínguez and Kolter.

Nongenetic individuality in the host-phage interaction.

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Sivan Pearl

2008-05-01

Full Text Available Isogenic bacteria can exhibit a range of phenotypes, even in homogeneous environmental conditions. Such nongenetic individuality has been observed in a wide range of biological processes, including differentiation and stress response. A striking example is the heterogeneous response of bacteria to antibiotics, whereby a small fraction of drug-sensitive bacteria can persist under extensive antibiotic treatments. We have previously shown that persistent bacteria enter a phenotypic state, identified by slow growth or dormancy, which protects them from the lethal action of antibiotics. Here, we studied the effect of persistence on the interaction between Escherichia coli and phage lambda. We used long-term time-lapse microscopy to follow the expression of green fluorescent protein (GFP under the phage lytic promoter, as well as cellular fate, in single infected bacteria. Intriguingly, we found that, whereas persistent bacteria are protected from prophage induction, they are not protected from lytic infection. Quantitative analysis of gene expression reveals that the expression of lytic genes is suppressed in persistent bacteria. However, when persistent bacteria switch to normal growth, the infecting phage resumes the process of gene expression, ultimately causing cell lysis. Using mathematical models for these two host-phage interactions, we found that the bacteria's nongenetic individuality can significantly affect the population dynamics, and might be relevant for understanding the coevolution of bacterial hosts and phages.

The detection of Salmonella typhimurium on shell eggs using a phage-based biosensor

Science.gov (United States)

Chai, Yating; Li, Suiqiong; Horikawa, Shin; Shen, Wen; Park, Mi-Kyung; Vodyanoy, Vitaly J.; Chin, Bryan A.

2011-06-01

This paper presents the direct detection of Salmonella typhimurium on shell eggs using a phage-based magnetoelastic (ME) biosensor. The ME biosensor consists of a ME resonator as the sensor platform and E2 phage as the biorecognition element that is genetically engineered to specifically bind with Salmonella typhimurium. The ME biosensor, which is a wireless sensor, vibrates with a characteristic resonant frequency under an externally applied magnetic field. Multiple sensors can easily be remotely monitored. Multiple measurement and control sensors were placed on the shell eggs contaminated by Salmonella typhimurium solutions with different known concentrations. The resonant frequency of sensors before and after the exposure to the spiked shell eggs was measured. The frequency shift of the measurement sensors was significantly different than the control sensors indicating Salmonella contamination. Scanning electron microscopy was used to confirm binding of Salmonella to the sensor surface and the resulting frequency shift results.

DNA damage and mutagenesis of lambda phage induced by gamma-rays

International Nuclear Information System (INIS)

Bertram, Heidi

1988-01-01

Lambda phage DNA was gamma irradiated in aqueous solution and strand breakage determined. Twice as much minor structural damage per lethal hit was found in this DNA compared with DNA from irradiated phage suspensions. The in vitro irradiated DNA was repackaged into infectious particles. Induction of mutations in the cI or cII cistron was scored using SOS-induced host cells. In vitro prepared particles were found to have second-order kinetics for mutagenesis induced by gamma rays indicating two pre-mutational events were necessary to produce a mutation, but bacteria-free phage suspensions ('lys-phage') showed single hit kinetics for mutagenesis after irradiation. Increase in the mutation rate in the phage particles was mainly due to minor lesions, i.e. ssb, als and unidentified base damage. In lys-phage, mutagenesis might be enhanced by clustered DNA damage - configuration not existing in pack-phage. Loss of infectivity was analysed in comparison with structural damage. All lesions contributed to biological inactivation. Minor lesions were tolerated by lambda phage to a limited extent. Major lesions (e.g. dsb) contributed most to infectivity loss and were considered lethal events. (U.K.)

Changes of the Specific Infectivity of Tracer Phages during Transport in Porous Media.

Science.gov (United States)

Ghanem, Nawras; Trost, Manuel; Sánchez Fontanet, Laura; Harms, Hauke; Chatzinotas, Antonis; Wick, Lukas Y

2018-03-20

Phages (i.e., viruses infecting bacteria) are considered to be good indicators and tracers for fecal pollution, hydraulic flow, or colloidal transport in the subsurface. They are typically quantified as total virus particles (VLP) or plaque forming units (PFU) of infectious phages. As transport may lead to phage deactivation, VLP quantification can overestimate the number of infectious phages. In contrast, PFU counts may underestimate the transport of total virus particles. Using PFU and tunable resistive pulse sensing-based counting for active and total phages, respectively, we quantified the effect of transport through laboratory percolation columns on the specific infectivity (SI). The SI is defined by the ratio of total VLP to PFU and is a measure for the minimum particle numbers needed to create a single infection. Transport of three marine tracer phages and the coli-phage (T4) was described by colloidal filtration theory. We found that apparent collision efficiencies of active and total phages differed. Depending on the phage properties (e.g., morphology or hydrophobicity), passage through a porous medium led to either an increasing or decreasing SI of effluent phages. Our data suggest that both phage mass recovery and the SI should be considered in quantitative phage tracer experiments.

Primary isolation strain determines both phage type and receptors recognised by Campylobacter jejuni bacteriophages.

Directory of Open Access Journals (Sweden)

Martine C Holst Sørensen

Full Text Available In this study we isolated novel bacteriophages, infecting the zoonotic bacterium Campylobacter jejuni. These phages may be used in phage therapy of C. jejuni colonized poultry to prevent spreading of the bacteria to meat products causing disease in humans. Many C. jejuni phages have been isolated using NCTC12662 as the indicator strain, which may have biased the selection of phages. A large group of C. jejuni phages rely on the highly diverse capsular polysaccharide (CPS for infection and recent work identified the O-methyl phosphoramidate modification (MeOPN of CPS as a phage receptor. We therefore chose seven C. jejuni strains each expressing different CPS structures as indicator strains in a large screening for phages in samples collected from free-range poultry farms. Forty-three phages were isolated using C. jejuni NCTC12658, NCTC12662 and RM1221 as host strains and 20 distinct phages were identified based on host range analysis and genome restriction profiles. Most phages were isolated using C. jejuni strains NCTC12662 and RM1221 and interestingly phage genome size (140 kb vs. 190 kb, host range and morphological appearance correlated with the isolation strain. Thus, according to C. jejuni phage grouping, NCTC12662 and NCTC12658 selected for CP81-type phages, while RM1221 selected for CP220-type phages. Furthermore, using acapsular ∆kpsM mutants we demonstrated that phages isolated on NCTC12658 and NCTC12662 were dependent on the capsule for infection. In contrast, CP220-type phages isolated on RM1221 were unable to infect non-motile ∆motA mutants, hence requiring motility for successful infection. Hence, the primary phage isolation strain determines both phage type (CP81 or CP220 as well as receptors (CPS or flagella recognised by the isolated phages.

The Human Gut Phage Community and Its Implications for Health and Disease.

Science.gov (United States)

Manrique, Pilar; Dills, Michael; Young, Mark J

2017-06-08

In this review, we assess our current understanding of the role of bacteriophages infecting the human gut bacterial community in health and disease. In general, bacteriophages contribute to the structure of their microbial communities by driving host and viral diversification, bacterial evolution, and by expanding the functional diversity of ecosystems. Gut bacteriophages are an ensemble of unique and shared phages in individuals, which encompass temperate phages found predominately as prophage in gut bacteria (prophage reservoir) and lytic phages. In healthy individuals, only a small fraction of the prophage reservoir is activated and found as extracellular phages. Phage community dysbiosis is characterized by a shift in the activated prophage community or an increase of lytic phages, and has been correlated with disease, suggesting that a proper balance between lysis and lysogeny is needed to maintain health. Consequently, the concept of microbial dysbiosis might be extended to the phage component of the microbiome as well. Understanding the dynamics and mechanisms to restore balance after dysbiosis is an active area of research. The use of phage transplants to re-establish health suggests that phages can be used as disease treatment. Such advances represent milestones in our understanding of gut phages in human health and should fuel research on their role in health and disease.

A CRISPR-Cas9 System for Genetic Engineering of Filamentous Fungi

DEFF Research Database (Denmark)

Nødvig, Christina Spuur; Nielsen, Jakob Blæsbjerg; Kogle, Martin Engelhard

2015-01-01

there is a demand for developing versatile methods that can be used to genetically manipulate non-model filamentous fungi. To facilitate this, we have developed a CRISPR-Cas9 based system adapted for use in filamentous fungi. The system is simple and versatile, as RNA guided mutagenesis can be achieved...... by transforming a target fungus with a single plasmid. The system currently contains four CRISPR- Cas9 vectors, which are equipped with commonly used fungal markers allowing for selection in a broad range of fungi. Moreover, we have developed a script that allows identification of protospacers that target gene...... used our CRISPR Cas9 system to generate a strain that contains an AACU_pyrG marker and demonstrated that the resulting strain can be used for iterative gene targeting....

Molecular Characterization of Three Lactobacillus delbrueckii subsp. bulgaricus Phages

OpenAIRE

Casey, Eoghan; Mahony, Jennifer; O'Connell-Motherway, Mary; Bottacini, Francesca; Cornelissen, Anneleen; Neve, Horst; Heller, Knut J.; Noben, Jean-Paul; Dal Bello, Fabio; van Sinderen, Douwe

2014-01-01

In this study, three phages infecting Lactobacillus delbrueckii subsp. bulgaricus, named Ld3, Ld17, and Ld25A, were isolated from whey samples obtained from various industrial fermentations. These phages were further characterized in a multifaceted approach: (i) biological and physical characterization through host range analysis and electron microscopy; (ii) genetic assessment through genome analysis; (iii) mass spectrometry analysis of the structural components of the phages; and (iv), for ...

Phage therapy in the food industry.

Science.gov (United States)

Endersen, Lorraine; O'Mahony, Jim; Hill, Colin; Ross, R Paul; McAuliffe, Olivia; Coffey, Aidan

2014-01-01

Despite advances in modern technologies, the food industry is continuously challenged with the threat of microbial contamination. The overuse of antibiotics has further escalated this problem, resulting in the increasing emergence of antibiotic-resistant foodborne pathogens. Efforts to develop new methods for controlling microbial contamination in food and the food processing environment are extremely important. Accordingly, bacteriophages (phages) and their derivatives have emerged as novel, viable, and safe options for the prevention, treatment, and/or eradication of these contaminants in a range of foods and food processing environments. Whole phages, modified phages, and their derivatives are discussed in terms of current uses and future potential as antimicrobials in the traditional farm-to-fork context, encompassing areas such as primary production, postharvest processing, biosanitation, and biodetection. The review also presents some safety concerns to ensure safe and effective exploitation of bacteriophages in the future.

THz field engineering in two-color femtosecond filaments using chirped and delayed laser pulses

Science.gov (United States)

Nguyen, A.; González de Alaiza Martínez, P.; Thiele, I.; Skupin, S.; Bergé, L.

2018-03-01

We numerically study the influence of chirping and delaying several ionizing two-color light pulses in order to engineer terahertz (THz) wave generation in air. By means of comprehensive 3D simulations, it is shown that two chirped pulses can increase the THz yield when they are separated by a suitable time delay for the same laser energy in focused propagation geometry. To interpret these results, the local current theory is revisited and we propose an easy, accessible all-optical criterion that predicts the laser-to-THz conversion efficiencies given any input laser spectrum. In the filamentation regime, numerical simulations display evidence that a chirped pulse is able to produce more THz radiation due to propagation effects, which maintain the two colors of the laser field more efficiently coupled over long distances. A large delay between two pulses promotes multi-peaked THz spectra as well as conversion efficiencies above 10‑4.

Tales of diversity: Genomic and morphological characteristics of forty-six Arthrobacter phages.

Directory of Open Access Journals (Sweden)

Karen K Klyczek

Full Text Available The vast bacteriophage population harbors an immense reservoir of genetic information. Almost 2000 phage genomes have been sequenced from phages infecting hosts in the phylum Actinobacteria, and analysis of these genomes reveals substantial diversity, pervasive mosaicism, and novel mechanisms for phage replication and lysogeny. Here, we describe the isolation and genomic characterization of 46 phages from environmental samples at various geographic locations in the U.S. infecting a single Arthrobacter sp. strain. These phages include representatives of all three virion morphologies, and Jasmine is the first sequenced podovirus of an actinobacterial host. The phages also span considerable sequence diversity, and can be grouped into 10 clusters according to their nucleotide diversity, and two singletons each with no close relatives. However, the clusters/singletons appear to be genomically well separated from each other, and relatively few genes are shared between clusters. Genome size varies from among the smallest of siphoviral phages (15,319 bp to over 70 kbp, and G+C contents range from 45-68%, compared to 63.4% for the host genome. Although temperate phages are common among other actinobacterial hosts, these Arthrobacter phages are primarily lytic, and only the singleton Galaxy is likely temperate.

Tales of diversity: Genomic and morphological characteristics of forty-six Arthrobacter phages.

Science.gov (United States)

Klyczek, Karen K; Bonilla, J Alfred; Jacobs-Sera, Deborah; Adair, Tamarah L; Afram, Patricia; Allen, Katherine G; Archambault, Megan L; Aziz, Rahat M; Bagnasco, Filippa G; Ball, Sarah L; Barrett, Natalie A; Benjamin, Robert C; Blasi, Christopher J; Borst, Katherine; Braun, Mary A; Broomell, Haley; Brown, Conner B; Brynell, Zachary S; Bue, Ashley B; Burke, Sydney O; Casazza, William; Cautela, Julia A; Chen, Kevin; Chimalakonda, Nitish S; Chudoff, Dylan; Connor, Jade A; Cross, Trevor S; Curtis, Kyra N; Dahlke, Jessica A; Deaton, Bethany M; Degroote, Sarah J; DeNigris, Danielle M; DeRuff, Katherine C; Dolan, Milan; Dunbar, David; Egan, Marisa S; Evans, Daniel R; Fahnestock, Abby K; Farooq, Amal; Finn, Garrett; Fratus, Christopher R; Gaffney, Bobby L; Garlena, Rebecca A; Garrigan, Kelly E; Gibbon, Bryan C; Goedde, Michael A; Guerrero Bustamante, Carlos A; Harrison, Melinda; Hartwell, Megan C; Heckman, Emily L; Huang, Jennifer; Hughes, Lee E; Hyduchak, Kathryn M; Jacob, Aswathi E; Kaku, Machika; Karstens, Allen W; Kenna, Margaret A; Khetarpal, Susheel; King, Rodney A; Kobokovich, Amanda L; Kolev, Hannah; Konde, Sai A; Kriese, Elizabeth; Lamey, Morgan E; Lantz, Carter N; Lapin, Jonathan S; Lawson, Temiloluwa O; Lee, In Young; Lee, Scott M; Lee-Soety, Julia Y; Lehmann, Emily M; London, Shawn C; Lopez, A Javier; Lynch, Kelly C; Mageeney, Catherine M; Martynyuk, Tetyana; Mathew, Kevin J; Mavrich, Travis N; McDaniel, Christopher M; McDonald, Hannah; McManus, C Joel; Medrano, Jessica E; Mele, Francis E; Menninger, Jennifer E; Miller, Sierra N; Minick, Josephine E; Nabua, Courtney T; Napoli, Caroline K; Nkangabwa, Martha; Oates, Elizabeth A; Ott, Cassandra T; Pellerino, Sarah K; Pinamont, William J; Pirnie, Ross T; Pizzorno, Marie C; Plautz, Emilee J; Pope, Welkin H; Pruett, Katelyn M; Rickstrew, Gabbi; Rimple, Patrick A; Rinehart, Claire A; Robinson, Kayla M; Rose, Victoria A; Russell, Daniel A; Schick, Amelia M; Schlossman, Julia; Schneider, Victoria M; Sells, Chloe A; Sieker, Jeremy W; Silva, Morgan P; Silvi, Marissa M; Simon, Stephanie E; Staples, Amanda K; Steed, Isabelle L; Stowe, Emily L; Stueven, Noah A; Swartz, Porter T; Sweet, Emma A; Sweetman, Abigail T; Tender, Corrina; Terry, Katrina; Thomas, Chrystal; Thomas, Daniel S; Thompson, Allison R; Vanderveen, Lorianna; Varma, Rohan; Vaught, Hannah L; Vo, Quynh D; Vonberg, Zachary T; Ware, Vassie C; Warrad, Yasmene M; Wathen, Kaitlyn E; Weinstein, Jonathan L; Wyper, Jacqueline F; Yankauskas, Jakob R; Zhang, Christine; Hatfull, Graham F

2017-01-01

The vast bacteriophage population harbors an immense reservoir of genetic information. Almost 2000 phage genomes have been sequenced from phages infecting hosts in the phylum Actinobacteria, and analysis of these genomes reveals substantial diversity, pervasive mosaicism, and novel mechanisms for phage replication and lysogeny. Here, we describe the isolation and genomic characterization of 46 phages from environmental samples at various geographic locations in the U.S. infecting a single Arthrobacter sp. strain. These phages include representatives of all three virion morphologies, and Jasmine is the first sequenced podovirus of an actinobacterial host. The phages also span considerable sequence diversity, and can be grouped into 10 clusters according to their nucleotide diversity, and two singletons each with no close relatives. However, the clusters/singletons appear to be genomically well separated from each other, and relatively few genes are shared between clusters. Genome size varies from among the smallest of siphoviral phages (15,319 bp) to over 70 kbp, and G+C contents range from 45-68%, compared to 63.4% for the host genome. Although temperate phages are common among other actinobacterial hosts, these Arthrobacter phages are primarily lytic, and only the singleton Galaxy is likely temperate.

Bacteriophage populations

International Nuclear Information System (INIS)

Klieve, A.V.; Gilbert, R.A.

2005-01-01

Bacteriophages are ubiquitous to the rumen ecosystem; they have a role in nitrogen metabolism through bacterial lysis in the rumen, they may help to regulate bacterial population densities, be an agent for genetic exchange and be of use in biocontrol of bacterial populations through phage therapy. In Chapter 2.1, classical methodologies to enable the isolation, enumeration, storage and morphological characterization of phages were presented. In addition to these classic procedures, molecular biological techniques have resulted in a range of methodologies to investigate the type, topology and size of phage nucleic acids, to fingerprint individual phage strains and to create a profile of ruminal phage populations. Different phage families possess all the currently identified combinations of double-stranded or single-stranded RNA or DNA and may also possess unusual bases such as 5-hydroxymethylcytosine (found in T-even phage) or 5- hydroxymethyluracil and uracil in place of thymidine. In all morphological groups of phage except the filamentous phages, the nucleic acid is contained within a head or polyhedral structure, predominantly composed of protein. Filamentous phages have their nucleic acid contained inside the helical filament, occupying much of its length. Many of the procedures used with phage nucleic acids and double-stranded (ds) DNA, in particular, are not specific to ruminal phages but are the same as in other areas where nucleic acids are investigated and are covered elsewhere in the literature and this chapter. Most applications with rumen phages are similar to those reported for phages of non-ruminal bacteria and are covered in general texts such as Maniatis et al. In this chapter, we will concentrate on aspects of methodology as they relate to ruminal phages

Ciprofloxacin and Trimethoprim Cause Phage Induction and Virulence Modulation in Staphylococcus aureus

Science.gov (United States)

Goerke, Christiane; Köller, Johanna; Wolz, Christiane

2006-01-01

In Staphylococcus aureus strains of human origin, phages which integrate into the chromosomal gene coding for β-hemolysin (hlb) are widely distributed. Most of them encode accessory virulence determinants such as staphylokinase (sak) or enterotoxins. Here, we analyzed the effects of ciprofloxacin and trimethoprim on phage induction and expression of phage-encoded virulence factors by using isolates from patients with cystic fibrosis for which the induction of hlb-converting phages was demonstrated in vivo (C. Goerke, S. Matias y Papenberg, S. Dasbach, K. Dietz, R. Ziebach, B. C. Kahl, and C. Wolz, J. Infect. Dis. 189:724-734, 2004) as well as a φ13 lysogen of phage-cured strain 8325-4. Treatment of lysogens with subinhibitory concentrations of either antibiotic resulted in (i) delysogenization of strains resembling the isolates picked up after chronic lung infection and (ii) replication of phages in the bacterial host in a dose-dependent manner. Ciprofloxacin treatment resulted in enhanced recA transcription, indicating involvement of the SOS response in phage mobilization. Induction of φ13 was linked to elevated expression of the phage-encoded virulence gene sak, chiefly due to the activation of latent phage promoters. In summary, we could show the induction of hlb-converting phages and a subsequent virulence modulation of the host bacterium by ciprofloxacin and trimethoprim. PMID:16377683

Inhaled phage therapy: a promising and challenging approach to treat bacterial respiratory infections.

Science.gov (United States)

Bodier-Montagutelli, Elsa; Morello, Eric; L'Hostis, Guillaume; Guillon, Antoine; Dalloneau, Emilie; Respaud, Renaud; Pallaoro, Nikita; Blois, Hélène; Vecellio, Laurent; Gabard, Jérôme; Heuzé-Vourc'h, Nathalie

2017-08-01

Bacterial respiratory tract infections (RTIs) are increasingly difficult to treat due to evolving antibiotic resistance. In this context, bacteriophages (or phages) are part of the foreseen alternatives or combination therapies. Delivering phages through the airways seems more relevant to accumulate these natural antibacterial viruses in proximity to their bacterial host, within the infectious site. Areas covered: This review addresses the potential of phage therapy to treat RTIs and discusses preclinical and clinical results of phages administration in this context. Recent phage formulation and aerosolization attempts are also reviewed, raising technical challenges to achieve efficient pulmonary deposition via inhalation. Expert opinion: Overall, the inhalation of phages as antibacterial treatment seems both clinically relevant and technically feasible. Several crucial points still need to be investigated, such as phage product pharmacokinetics and immunogenicity. Furthermore, given phage-specific features, appropriate regulatory and manufacturing guidelines will need to be defined. Finally, randomized controlled clinical trials should be carried out to establish phage therapy's clinical positioning in the antimicrobial arsenal against RTIs.

Dual-functioning peptides discovered by phage display increase the magnitude and specificity of BMSC attachment to mineralized biomaterials.

Science.gov (United States)

Ramaraju, Harsha; Miller, Sharon J; Kohn, David H

2017-07-01

Design of biomaterials for cell-based therapies requires presentation of specific physical and chemical cues to cells, analogous to cues provided by native extracellular matrices (ECM). We previously identified a peptide sequence with high affinity towards apatite (VTKHLNQISQSY, VTK) using phage display. The aims of this study were to identify a human MSC-specific peptide sequence through phage display, combine it with the apatite-specific sequence, and verify the specificity of the combined dual-functioning peptide to both apatite and human bone marrow stromal cells. In this study, a combinatorial phage display identified the cell binding sequence (DPIYALSWSGMA, DPI) which was combined with the mineral binding sequence to generate the dual peptide DPI-VTK. DPI-VTK demonstrated significantly greater binding affinity (1/K D ) to apatite surfaces compared to VTK, phosphorylated VTK (VTK phos ), DPI-VTK phos , RGD-VTK, and peptide-free apatite surfaces (p biomaterial surfaces and subsequently increase cell proliferation and differentiation. These new peptides expand biomaterial design methodology for cell-based regeneration of bone defects. This strategy of combining cell and material binding phage display derived peptides is broadly applicable to a variety of systems requiring targeted adhesion of specific cell populations, and may be generalized to the engineering of any adhesion surface. Copyright © 2017 Elsevier Ltd. All rights reserved.

Differential screening of phage-ab libraries by oligonucleotide microarray technology.

Directory of Open Access Journals (Sweden)

Paolo Monaci

Full Text Available A novel and efficient tagArray technology was developed that allows rapid identification of antibodies which bind to receptors with a specific expression profile, in the absence of biological information. This method is based on the cloning of a specific, short nucleotide sequence (tag in the phagemid coding for each phage-displayed antibody fragment (phage-Ab present in a library. In order to set up and validate the method we identified about 10,000 different phage-Abs binding to receptors expressed in their native form on the cell surface (10 k Membranome collection and tagged each individual phage-Ab. The frequency of each phage-Ab in a given population can at this point be inferred by measuring the frequency of its associated tag sequence through standard DNA hybridization methods. Using tiny amounts of biological samples we identified phage-Abs binding to receptors preferentially expressed on primary tumor cells rather than on cells obtained from matched normal tissues. These antibodies inhibited cell proliferation in vitro and tumor development in vivo, thus representing therapeutic lead candidates.

Genomics of phages with therapeutic potential

DEFF Research Database (Denmark)

Zschach, Henrike

Bacteriophages, viruses that prey on bacteria, have been applied since the 1920’s to treat and prevent bacterial infection. After the discovery of antibiotics, this route was however largely abandoned. Now, with antimicrobial resistance in human-pathogenic bacteria on the rise and a dire need...... for alternatives, phage therapy once again takes center stage. Phage therapy holds the promise of substantial benefits both from the economic as well as the public health perspective but also holds distinct challenges. The aim of this PhD was to address how bioinformatics tools, specifically genomics...... and mathematical modelling, can be applied to move the field towards a future of actual phage therapy in humans. It is composed of three related research projects. The first part of this thesis is an introduction to various topics and methods relevant to the research projects that jointedly make up this Ph...

Morphology and rheology in filamentous cultivations.

Science.gov (United States)

Wucherpfennig, T; Kiep, K A; Driouch, H; Wittmann, C; Krull, R

2010-01-01

Because of their metabolic diversity, high production capacity, secretion efficiency, and capability of carrying out posttranslational modifications, filamentous fungi are widely exploited as efficient cell factories in the production of metabolites, bioactive substances, and native or heterologous proteins, respectively. There is, however, a complex relationship between the morphology of these microorganisms, transport phenomena, the viscosity of the cultivation broth, and related productivity. The morphological characteristics vary between freely dispersed mycelia and distinct pellets of aggregated biomass, every growth form having a distinct influence on broth rheology. Hence, the advantages and disadvantages for mycelial or pellet cultivation have to be balanced out carefully. Because of the still inadequate understanding of the morphogenesis of filamentous microorganisms, fungal morphology is often a bottleneck of productivity in industrial production. To obtain an optimized production process, it is of great importance to gain a better understanding of the molecular and cell biology of these microorganisms as well as the relevant approaches in biochemical engineering. In this chapter, morphology and growth of filamentous fungi are described, with special attention given to specific problems as they arise from fungal growth forms; growth and mass transfer in fungal biopellets are discussed as an example. To emphasize the importance of the flow behavior of filamentous cultivation broths, an introduction to rheology is also given, reviewing important rheological models and recent studies concerning rheological parameters. Furthermore, current knowledge on morphology and productivity in relation to the environom is outlined in the last section of this review. Copyright 2010 Elsevier Inc. All rights reserved.

Identification of the host determinant of two prolate-headed phages infecting lactococcus lactis

International Nuclear Information System (INIS)

Stuer-Lauridsen, Birgitte; Janzen, Thomas; Schnabl, Jannie; Johansen, Eric

2003-01-01

A gene responsible for host determination was identified in two prolate-headed bacteriophages of the c2 species infecting strains of Lactococcus lactis. The identification of the host determinant gene was based on low DNA sequence homology in a specific open reading frame (ORF) between prolate-headed phages with different host ranges. When a host carrying this ORF from one phage on a plasmid was infected with another phage, we obtained phages with an altered host range at a frequency of 10 -6 to 10 -7 . Sequencing of phage DNA originating from 10 independent single plaques confirmed that a genetic recombination had taken place at different positions between the ORF on the plasmid and the infecting phage. The adsorption of the recombinant phages to their bacterial hosts had also changed to match the phage origin of the ORF. Consequently, it is concluded that this ORF codes for the host range determinant

Phage therapy against Enterococcus faecalis in dental root canals

Directory of Open Access Journals (Sweden)

Leron Khalifa

2016-09-01

Full Text Available Antibiotic resistance is an ever-growing problem faced by all major sectors of health care, including dentistry. Recurrent infections related to multidrug-resistant bacteria such as methicillin-resistant Staphylococcus aureus, carbapenem-resistant Enterobacteriaceae, and vancomycin-resistant enterococci (VRE in hospitals are untreatable and question the effectiveness of notable drugs. Two major reasons for these recurrent infections are acquired antibiotic resistance genes and biofilm formation. None of the traditionally known effective techniques have been able to efficiently resolve these issues. Hence, development of a highly effective antibacterial practice has become inevitable. One example of a hard-to-eradicate pathogen in dentistry is Enterococcus faecalis, which is one of the most common threats observed in recurrent root canal treatment failures, of which the most problematic to treat are its biofilm-forming VRE strains. An effective response against such infections could be the use of bacteriophages (phages. Phage therapy was found to be highly effective against biofilm and multidrug-resistant bacteria and has other advantages like ease of isolation and possibilities for genetic manipulations. The potential of phage therapy in dentistry, in particular against E. faecalis biofilms in root canals, is almost unexplored. Here we review the efforts to develop phage therapy against biofilms. We also focus on the phages isolated against E. faecalis and discuss the possibility of using phages against E. faecalis biofilm in root canals.

Recombinant lambda-phage nanobioparticles for tumor therapy in mice models.

Science.gov (United States)

Ghaemi, Amir; Soleimanjahi, Hoorieh; Gill, Pooria; Hassan, Zuhair; Jahromi, Soodeh Razeghi M; Roohvand, Farzin

2010-05-12

Lambda phages have considerable potential as gene delivery vehicles due to their genetic tractability, low cost, safety and physical characteristics in comparison to other nanocarriers and gene porters. Little is known concerning lambda phage-mediated gene transfer and expression in mammalian hosts. We therefore performed experiments to evaluate lambda-ZAP bacteriophage-mediated gene transfer and expression in vitro. For this purpose, we constructed recombinant lambda-phage nanobioparticles containing a mammalian expression cassette encoding enhanced green fluorescent protein (EGFP) and E7 gene of human papillomavirus type 16 (lambda-HPV-16 E7) using Lambda ZAP- CMV XR vector. Four cell lines (COS-7, CHO, TC-1 and HEK-239) were transduced with the nanobioparticles. We also characterized the therapeutic anti-tumor effects of the recombinant lambda-HPV-16 E7 phage in C57BL/6 tumor mice model as a cancer vaccine. Obtained results showed that delivery and expression of these genes in fibroblastic cells (COS-7 and CHO) are more efficient than epithelial cells (TC-1 and HEK-239) using these nanobioparticles. Despite the same phage M.O.I entry, the internalizing titers of COS-7 and CHO cells were more than TC-1 and HEK-293 cells, respectively. Mice vaccinated with lambda-HPV-16 E7 are able to generate potent therapeutic antitumor effects against challenge with E7- expressing tumor cell line, TC-1 compared to group treated with the wild phage. The results demonstrated that the recombinant lambda-phages, due to their capabilities in transducing mammalian cells, can also be considered in design and construction of novel and safe phage-based nanomedicines.

The versatility of hot-filament activated chemical vapor deposition

International Nuclear Information System (INIS)

Schaefer, Lothar; Hoefer, Markus; Kroeger, Roland

2006-01-01

In the field of activated chemical vapor deposition (CVD) of polycrystalline diamond films, hot-filament activation (HF-CVD) is widely used for applications where large deposition areas are needed or three-dimensional substrates have to be coated. We have developed processes for the deposition of conductive, boron-doped diamond films as well as for tribological crystalline diamond coatings on deposition areas up to 50 cm x 100 cm. Such multi-filament processes are used to produce diamond electrodes for advanced electrochemical processes or large batches of diamond-coated tools and parts, respectively. These processes demonstrate the high degree of uniformity and reproducibility of hot-filament CVD. The usability of hot-filament CVD for diamond deposition on three-dimensional substrates is well known for CVD diamond shaft tools. We also develop interior diamond coatings for drawing dies, nozzles, and thread guides. Hot-filament CVD also enables the deposition of diamond film modifications with tailored properties. In order to adjust the surface topography to specific applications, we apply processes for smooth, fine-grained or textured diamond films for cutting tools and tribological applications. Rough diamond is employed for grinding applications. Multilayers of fine-grained and coarse-grained diamond have been developed, showing increased shock resistance due to reduced crack propagation. Hot-filament CVD is also used for in situ deposition of carbide coatings and diamond-carbide composites, and the deposition of non-diamond, silicon-based films. These coatings are suitable as diffusion barriers and are also applied for adhesion and stress engineering and for semiconductor applications, respectively

Manipulating or superseding host recombination functions: a dilemma that shapes phage evolvability.

Directory of Open Access Journals (Sweden)

Louis-Marie Bobay

Full Text Available Phages, like many parasites, tend to have small genomes and may encode autonomous functions or manipulate those of their hosts'. Recombination functions are essential for phage replication and diversification. They are also nearly ubiquitous in bacteria. The E. coli genome encodes many copies of an octamer (Chi motif that upon recognition by RecBCD favors repair of double strand breaks by homologous recombination. This might allow self from non-self discrimination because RecBCD degrades DNA lacking Chi. Bacteriophage Lambda, an E. coli parasite, lacks Chi motifs, but escapes degradation by inhibiting RecBCD and encoding its own autonomous recombination machinery. We found that only half of 275 lambdoid genomes encode recombinases, the remaining relying on the host's machinery. Unexpectedly, we found that some lambdoid phages contain extremely high numbers of Chi motifs concentrated between the phage origin of replication and the packaging site. This suggests a tight association between replication, packaging and RecBCD-mediated recombination in these phages. Indeed, phages lacking recombinases strongly over-represent Chi motifs. Conversely, phages encoding recombinases and inhibiting host recombination machinery select for the absence of Chi motifs. Host and phage recombinases use different mechanisms and the latter are more tolerant to sequence divergence. Accordingly, we show that phages encoding their own recombination machinery have more mosaic genomes resulting from recent recombination events and have more diverse gene repertoires, i.e. larger pan genomes. We discuss the costs and benefits of superseding or manipulating host recombination functions and how this decision shapes phage genome structure and evolvability.

Complete Genome Sequences of 44 Arthrobacter Phages.

Science.gov (United States)

Klyczek, Karen K; Jacobs-Sera, Deborah; Adair, Tamarah L; Adams, Sandra D; Ball, Sarah L; Benjamin, Robert C; Bonilla, J Alfred; Breitenberger, Caroline A; Daniels, Charles J; Gaffney, Bobby L; Harrison, Melinda; Hughes, Lee E; King, Rodney A; Krukonis, Gregory P; Lopez, A Javier; Monsen-Collar, Kirsten; Pizzorno, Marie C; Rinehart, Claire A; Staples, Amanda K; Stowe, Emily L; Garlena, Rebecca A; Russell, Daniel A; Cresawn, Steven G; Pope, Welkin H; Hatfull, Graham F

2018-02-01

We report here the complete genome sequences of 44 phages infecting Arthrobacter sp. strain ATCC 21022. These phages have double-stranded DNA genomes with sizes ranging from 15,680 to 70,707 bp and G+C contents from 45.1% to 68.5%. All three tail types (belonging to the families Siphoviridae , Myoviridae , and Podoviridae ) are represented. Copyright © 2018 Klyczek et al.

Significance of phage-host interactions for biocontrol of Campylobacter jejuni in food

DEFF Research Database (Denmark)

Athina, Zampara; Sørensen, Martine Camilla Holst; Elsser-Gravesen, Anne

2017-01-01

Poultry meat is the main source of Campylobacter jejuni foodborne disease. Currently, no effective control measures prevent C. jejuni from contaminating poultry meat. However, post-harvest phage treatment is a promising biocontrol strategy that has not yet been explored. Here we identified phages....... A thorough understanding of phage-host interactions is prerequisite to further advance phage application as a post-harvest biocontrol strategy against C. jejuni....

Genome Sequences of Gordonia Phages BaxterFox, Kita, Nymphadora, and Yeezy

OpenAIRE

Pope, Welkin H.; Bandla, Sharanya; Colbert, Alexandra K.; Eichinger, Fiona G.; Gamburg, Michelle B.; Horiates, Stavroula G.; Jamison, Jerrica M.; Julian, Dana R.; Moore, Whitney A.; Murthy, Pranav; Powell, Meghan C.; Smith, Sydney V.; Mezghani, Nadia; Milliken, Katherine A.; Thompson, Paige K.

2016-01-01

Gordonia phages BaxterFox, Kita, Nymphadora, and Yeezy are newly characterized phages of Gordonia terrae, isolated from soil samples in Pittsburgh, Pennsylvania. These phages have genome lengths between 50,346 and 53,717?bp, and encode on average 84 predicted proteins. All have G+C content of 66.6%.

Effect of Bacteriophages on the Growth of Flavobacterium psychrophilum and Development of Phage-Resistant Strains

DEFF Research Database (Denmark)

Christiansen, Rói Hammershaimb; Madsen, Lone; Dalsgaard, Inger

2016-01-01

The controlling effect of single and multiple phages on the density of Flavobacterium psychrophilum at different initial multiplicity of infection (MOI) was assessed in batch cultures to explore the potential for phage-based treatment of this important fish pathogen. A high initial phage concentr......The controlling effect of single and multiple phages on the density of Flavobacterium psychrophilum at different initial multiplicity of infection (MOI) was assessed in batch cultures to explore the potential for phage-based treatment of this important fish pathogen. A high initial phage...... concentration (MOI = 0.3–4) was crucial for efficient viral lysis, resulting in a 104–105-fold reduction of phage-sensitive cells (both single phages and phage cocktails), which was maintained throughout the incubation (>10 days). Following cell lysis, regrowth of phage-resistant strains was examined...... and resistant strains were isolated for further characterization. The application of a mathematical model allowed simulation of phage-host interactions and resistance development, confirming indications from strain isolations that phage-sensitive strains dominated the regrowing population (>99.8 %) at low MOI...

Filamentary structures in dense plasma focus: Current filaments or vortex filaments?

Energy Technology Data Exchange (ETDEWEB)

Soto, Leopoldo, E-mail: lsoto@cchen.cl; Pavez, Cristian; Moreno, José [Comisión Chilena de Energía Nuclear, CCHEN, Casilla 188-D, Santiago (Chile); Center for Research and Applications in Plasma Physics and Pulsed Power, P4, Departamento de Ciencias Físicas, Facultad de Ciencias Exactas, Universidad Andrés Bello, República 220, Santiago (Chile); Castillo, Fermin [Universidad Nacional Autónoma de México, Cuernavaca, México (Mexico); Veloso, Felipe [Instituto de Física, Pontificia Universidad Católica de Chile, 7820436 Santiago (Chile); Auluck, S. K. H. [Bhabha Atomic Research Center, Mumbai 400 085 (India)

2014-07-15

Recent observations of an azimuthally distributed array of sub-millimeter size sources of fusion protons and correlation between extreme ultraviolet (XUV) images of filaments with neutron yield in PF-1000 plasma focus have re-kindled interest in their significance. These filaments have been described variously in literature as current filaments and vortex filaments, with very little experimental evidence in support of either nomenclature. This paper provides, for the first time, experimental observations of filaments on a table-top plasma focus device using three techniques: framing photography of visible self-luminosity from the plasma, schlieren photography, and interferometry. Quantitative evaluation of density profile of filaments from interferometry reveals that their radius closely agrees with the collision-less ion skin depth. This is a signature of relaxed state of a Hall fluid, which has significant mass flow with equipartition between kinetic and magnetic energy, supporting the “vortex filament” description. This interpretation is consistent with empirical evidence of an efficient energy concentration mechanism inferred from nuclear reaction yields.

Identifying Bacterial Immune Evasion Proteins Using Phage Display.

Science.gov (United States)

Fevre, Cindy; Scheepmaker, Lisette; Haas, Pieter-Jan

2017-01-01

Methods aimed at identification of immune evasion proteins are mainly rely on in silico prediction of sequence, structural homology to known evasion proteins or use a proteomics driven approach. Although proven successful these methods are limited by a low efficiency and or lack of functional identification. Here we describe a high-throughput genomic strategy to functionally identify bacterial immune evasion proteins using phage display technology. Genomic bacterial DNA is randomly fragmented and ligated into a phage display vector that is used to create a phage display library expressing bacterial secreted and membrane bound proteins. This library is used to select displayed bacterial secretome proteins that interact with host immune components.

Genetic characterization of ØVC8 lytic phage for Vibrio cholerae O1.

Science.gov (United States)

Solís-Sánchez, Alejandro; Hernández-Chiñas, Ulises; Navarro-Ocaña, Armando; De la Mora, Javier; Xicohtencatl-Cortes, Juan; Eslava-Campos, Carlos

2016-03-22

Epidemics and pandemics of cholera, a diarrheal disease, are attributed to Vibrio cholera serogroups O1 and O139. In recent years, specific lytic phages of V. cholera have been proposed to be important factors in the cyclic occurrence of cholera in endemic areas. However, the role and potential participation of lytic phages during long interepidemic periods of cholera in non-endemic regions have not yet been described. The purpose of this study was to isolate and characterize specific lytic phages of V. cholera O1 strains. Sixteen phages were isolated from wastewater samples collected at the Endhó Dam in Hidalgo State, Mexico, concentrated with PEG/NaCl, and purified by density gradient. The lytic activity of the purified phages was tested using different V. cholerae O1 and O139 strains. Phage morphology was visualized by transmission electron microscopy (TEM), and phage genome sequencing was performed using the Genome Analyzer IIx System. Genome assembly and bioinformatics analysis were performed using a set of high-throughput programs. Phage structural proteins were analyzed by mass spectrometry. Sixteen phages with lytic and lysogenic activity were isolated; only phage ØVC8 showed specific lytic activity against V. cholerae O1 strains. TEM images of ØVC8 revealed a phage with a short tail and an isometric head. The ØVC8 genome comprises linear double-stranded DNA of 39,422 bp with 50.8 % G + C. Of the 48 annotated ORFs, 16 exhibit homology with sequences of known function and several conserved domains. Bioinformatics analysis showed multiple conserved domains, including an Ig domain, suggesting that ØVC8 might adhere to different mucus substrates such as the human intestinal epithelium. The results suggest that ØVC8 genome utilize the "single-stranded cohesive ends" packaging strategy of the lambda-like group. The two structural proteins sequenced and analyzed are proteins of known function. ØVC8 is a lytic phage with specific activity against V. cholerae

Popping the cork: mechanisms of phage genome ejection

NARCIS (Netherlands)

Molineux, I.J.; Panja, D.

2013-01-01

Sixty years after Hershey and Chase showed that nucleic acid is the major component of phage particles that is ejected into cells, we still do not fully understand how the process occurs. Advances in electron microscopy have revealed the structure of the condensed DNA confined in a phage capsid, and

Filament Activation in Response to Magnetic Flux Emergence and Cancellation in Filament Channels

Science.gov (United States)

Li, Ting; Zhang, Jun; Ji, Haisheng

2015-06-01

We conducted a comparative analysis of two filaments that showed a quite different activation in response to the flux emergence within the filament channels. The observations from the Solar Dynamics Observatory (SDO) and Global Oscillation Network Group (GONG) were made to analyze the two filaments on 2013 August 17 - 20 (SOL2013-08-17) and September 29 (SOL2013-09-29). The first event showed that the main body of the filament was separated into two parts when an active region (AR) emerged with a maximum magnetic flux of about 6.4×1021 Mx underlying the filament. The close neighborhood and common direction of the bright threads in the filament and the open AR fan loops suggest a similar magnetic connectivity of these two flux systems. The equilibrium of the filament was not destroyed three days after the start of the emergence of the AR. To our knowledge, similar observations have never been reported before. In the second event, the emerging flux occurred nearby a barb of the filament with a maximum magnetic flux of 4.2×1020 Mx, about one order of magnitude lower than that of the first event. Two patches of parasitic polarity in the vicinity of the barb merged, then cancelled with nearby network fields. About 20 hours after the onset of the emergence, the filament erupted. Our findings imply that the location of emerging flux within the filament channel is probably crucial to filament evolution. If the flux emergence appears nearby the barbs, it is highly likely that the emerging flux and the filament magnetic fields will cancel, which may lead to the eruption of the filament. The comparison of the two events shows that the emergence of a small AR may still not be enough to disrupt the stability of a filament system, and the actual eruption only occurs after the flux cancellation sets in.

Lytic phages obscure the cost of antibiotic resistance in Escherichia coli.

Science.gov (United States)

Tazzyman, Samuel J; Hall, Alex R

2015-03-17

The long-term persistence of antibiotic-resistant bacteria depends on their fitness relative to other genotypes in the absence of drugs. Outside the laboratory, viruses that parasitize bacteria (phages) are ubiquitous, but costs of antibiotic resistance are typically studied in phage-free experimental conditions. We used a mathematical model and experiments with Escherichia coli to show that lytic phages strongly affect the incidence of antibiotic resistance in drug-free conditions. Under phage parasitism, the likelihood that antibiotic-resistant genetic backgrounds spread depends on their initial frequency, mutation rate and intrinsic growth rate relative to drug-susceptible genotypes, because these parameters determine relative rates of phage-resistance evolution on different genetic backgrounds. Moreover, the average cost of antibiotic resistance in terms of intrinsic growth in the antibiotic-free experimental environment was small relative to the benefits of an increased mutation rate in the presence of phages. This is consistent with our theoretical work indicating that, under phage selection, typical costs of antibiotic resistance can be outweighed by realistic increases in mutability if drug resistance and hypermutability are genetically linked, as is frequently observed in clinical isolates. This suggests the long-term distribution of antibiotic resistance depends on the relative rates at which different lineages adapt to other types of selection, which in the case of phage parasitism is probably extremely common, as well as costs of resistance inferred by classical in vitro methods.

Phage typing or CRISPR typing for epidemiological surveillance of Salmonella Typhimurium?

Science.gov (United States)

Mohammed, Manal

2017-11-07

Salmonella Typhimurium is the most dominant Salmonella serovar around the world. It is associated with foodborne gastroenteritis outbreaks but has recently been associated with invasive illness and deaths. Characterization of S. Typhimurium is therefore very crucial for epidemiological surveillance. Phage typing has been used for decades for subtyping of S. Typhimurium to determine the epidemiological relation among isolates. Recent studies however have suggested that high throughput clustered regular interspaced short palindromic repeats (CRISPR) typing has the potential to replace phage typing. This study aimed to determine the efficacy of high-throughput CRISPR typing over conventional phage typing in epidemiological surveillance and outbreak investigation of S. Typhimurium. In silico analysis of whole genome sequences (WGS) of well-documented phage types of S. Typhimurium reveals the presence of different CRISPR type among strains belong to the same phage type. Furthermore, different phage types of S. Typhimurium share identical CRISPR type. Interestingly, identical spacers were detected among outbreak and non-outbreak associated DT8 strains of S. Typhimurium. Therefore, CRISPR typing is not useful for the epidemiological surveillance and outbreak investigation of S. Typhimurium and phage typing, until it is replaced by WGS, is still the gold standard method for epidemiological surveillance of S. Typhimurium.

A fast method for large-scale isolation of phages from hospital ...

African Journals Online (AJOL)

This plaque-forming method could be adopted to isolate E. coli phage easily, rapidly and in large quantities. Among the 18 isolated E. coli phages, 10 of them had a broad host range in E. coli and warrant further study. Key words: Escherichia coli phages, large-scale isolation, drug resistance, biological properties.

Characterizing RecA-independent induction of Shiga toxin2-encoding phages by EDTA treatment.

Directory of Open Access Journals (Sweden)

Lejla Imamovic

Full Text Available BACKGROUND: The bacteriophage life cycle has an important role in Shiga toxin (Stx expression. The induction of Shiga toxin-encoding phages (Stx phages increases toxin production as a result of replication of the phage genome, and phage lysis of the host cell also provides a means of Stx toxin to exit the cell. Previous studies suggested that prophage induction might also occur in the absence of SOS response, independently of RecA. METHODOLOGY/PRINCIPAL FINDINGS: The influence of EDTA on RecA-independent Stx2 phage induction was assessed, in laboratory lysogens and in EHEC strains carrying Stx2 phages in their genome, by Real-Time PCR. RecA-independent mechanisms described for phage λ induction (RcsA and DsrA were not involved in Stx2 phage induction. In addition, mutations in the pathway for the stress response of the bacterial envelope to EDTA did not contribute to Stx2 phage induction. The effect of EDTA on Stx phage induction is due to its chelating properties, which was also confirmed by the use of citrate, another chelating agent. Our results indicate that EDTA affects Stx2 phage induction by disruption of the bacterial outer membrane due to chelation of Mg(2+. In all the conditions evaluated, the pH value had a decisive role in Stx2 phage induction. CONCLUSIONS/SIGNIFICANCE: Chelating agents, such as EDTA and citrate, induce Stx phages, which raises concerns due to their frequent use in food and pharmaceutical products. This study contributes to our understanding of the phenomenon of induction and release of Stx phages as an important factor in the pathogenicity of Shiga toxin-producing Escherichia coli (STEC and in the emergence of new pathogenic strains.

Prevalence, Host Range, and Comparative Genomic Analysis of Temperate Ochrobactrum Phages

Directory of Open Access Journals (Sweden)

Claudia Jäckel

2017-06-01

Full Text Available Ochrobactrum and Brucella are closely related bacteria that populate different habitats and differ in their pathogenic properties. Only little is known about mobile genetic elements in these genera which might be important for survival and virulence. Previous studies on Brucella lysogeny indicated that active phages are rare in this genus. To gain insight into the presence and nature of prophages in Ochrobactrum, temperate phages were isolated from various species and characterized in detail. In silico analyses disclosed numerous prophages in published Ochrobactrum genomes. Induction experiments showed that Ochrobactrum prophages can be induced by various stress factors and that some strains released phage particles even under non-induced conditions. Sixty percent of lysates prepared from 125 strains revealed lytic activity. The host range and DNA similarities of 19 phages belonging to the families Myoviridae, Siphoviridae, or Podoviridae were determined suggesting that they are highly diverse. Some phages showed relationship to the temperate Brucella inopinata phage BiPB01. The genomic sequences of the myovirus POA1180 (41,655 bp and podovirus POI1126 (60,065 bp were analyzed. Phage POA1180 is very similar to a prophage recently identified in a Brucella strain isolated from an exotic frog. The POA1180 genome contains genes which may confer resistance to chromate and the ability to take up sulfate. Phage POI1126 is related to podoviruses of Sinorhizobium meliloti (PCB5, Erwinia pyrifoliae (Pep14, and Burkholderia cenocepacia (BcepIL02 and almost identical to an unnamed plasmid of the Ochrobactrum intermedium strain LMG 3301. Further experiments revealed that the POI1126 prophage indeed replicates as an extrachromosomal element. The data demonstrate for the first time that active prophages are common in Ochrobactrum and suggest that atypical brucellae also may be a reservoir for temperate phages.

Stochasticity in the Expression of LamB and its Affect on λ phage Infection

Science.gov (United States)

Chapman, Emily; Wu, Xiao-Lun

2006-03-01

λ phage binds to E. Coli's lamB protein and injects its DNA into the cell. The phage quickly replicates and after a latent period the bacteria bursts, emitting mature phages. We developed a mathematical model based on the known physical events that occur when a λ phage infects an E.Coli cell. The results of these models predict that the bacteria and phage populations become extinct unless the parameters of the model are very finely tuned, which is untrue in the nature. The lamB protein is part of the maltose regulon and can be repressed to minimal levels when grown in the absence of inducer. Therefore, a cell that is not expressing any lamB protein at that moment is resistant against phage infection. We studied the dynamic relationship between λ phage and E. Coli when the concentration of phage greatly outnumbers the concentration of bacteria. We study how the stochasticity of the expression of lamB affects the percentage of cells that the λ phage infects. We show that even in the case when the maltose regulon is fully induced a percentage of cells continue to persist against phage infection.

Elastic deformation and failure in protein filament bundles: Atomistic simulations and coarse-grained modeling.

Science.gov (United States)

Hammond, Nathan A; Kamm, Roger D

2008-07-01

The synthetic peptide RAD16-II has shown promise in tissue engineering and drug delivery. It has been studied as a vehicle for cell delivery and controlled release of IGF-1 to repair infarcted cardiac tissue, and as a scaffold to promote capillary formation for an in vitro model of angiogenesis. The structure of RAD16-II is hierarchical, with monomers forming long beta-sheets that pair together to form filaments; filaments form bundles approximately 30-60 nm in diameter; branching networks of filament bundles form macroscopic gels. We investigate the mechanics of shearing between the two beta-sheets constituting one filament, and between cohered filaments of RAD16-II. This shear loading is found in filament bundle bending or in tensile loading of fibers composed of partial-length filaments. Molecular dynamics simulations show that time to failure is a stochastic function of applied shear stress, and that for a given loading time behavior is elastic for sufficiently small shear loads. We propose a coarse-grained model based on Langevin dynamics that matches molecular dynamics results and facilities extending simulations in space and time. The model treats a filament as an elastic string of particles, each having potential energy that is a periodic function of its position relative to the neighboring filament. With insight from these simulations, we discuss strategies for strengthening RAD16-II and similar materials.

Primary Isolation Strain Determines Both Phage Type and Receptors Recognised by Campylobacter jejuni Bacteriophages

DEFF Research Database (Denmark)

Sørensen, Martine C. Holst; Gencay, Yilmaz Emre; Birk, Tina

2015-01-01

were identified based on host range analysis and genome restriction profiles. Most phages were isolated using C. jejuni strains NCTC12662 and RM1221 and interestingly phage genome size (140 kb vs. 190 kb), host range and morphological appearance correlated with the isolation strain. Thus, according......In this study we isolated novel bacteriophages, infecting the zoonotic bacterium Campylobacter jejuni. These phages may be used in phage therapy of C. jejuni colonized poultry to prevent spreading of the bacteria to meat products causing disease in humans. Many C. jejuni phages have been isolated...... therefore chose seven C. jejuni strains each expressing different CPS structures as indicator strains in a large screening for phages in samples collected from free-range poultry farms. Forty-three phages were isolated using C. jejuni NCTC12658, NCTC12662 and RM1221 as host strains and 20 distinct phages...

Genome Sequences of Gordonia Phages BaxterFox, Kita, Nymphadora, and Yeezy.

Science.gov (United States)

Pope, Welkin H; Bandla, Sharanya; Colbert, Alexandra K; Eichinger, Fiona G; Gamburg, Michelle B; Horiates, Stavroula G; Jamison, Jerrica M; Julian, Dana R; Moore, Whitney A; Murthy, Pranav; Powell, Meghan C; Smith, Sydney V; Mezghani, Nadia; Milliken, Katherine A; Thompson, Paige K; Toner, Chelsea L; Ulbrich, Megan C; Furbee, Emily C; Grubb, Sarah R; Warner, Marcie H; Montgomery, Matthew T; Garlena, Rebecca A; Russell, Daniel A; Jacobs-Sera, Deborah; Hatfull, Graham F

2016-08-11

Gordonia phages BaxterFox, Kita, Nymphadora, and Yeezy are newly characterized phages of Gordonia terrae, isolated from soil samples in Pittsburgh, Pennsylvania. These phages have genome lengths between 50,346 and 53,717 bp, and encode on average 84 predicted proteins. All have G+C content of 66.6%. Copyright © 2016 Pope et al.

Diversity of phage infection types and associated terminology: the problem with 'Lytic or lysogenic'.

Science.gov (United States)

Hobbs, Zack; Abedon, Stephen T

2016-04-01

Bacteriophages, or phages, are viruses of members of domain Bacteria. These viruses play numerous roles in shaping the diversity of microbial communities, with impact differing depending on what infection strategies specific phages employ. From an applied perspective, these especially are communities containing undesired or pathogenic bacteria that can be modified through phage-mediated bacterial biocontrol, that is, through phage therapy. Here we seek to categorize phages in terms of their infection strategies as well as review or suggest more descriptive, accurate or distinguishing terminology. Categories can be differentiated in terms of (1) whether or not virion release occurs (productive infections versus lysogeny, pseudolysogeny and/or the phage carrier state), (2) the means of virion release (lytic versus chronic release) and (3) the degree to which phages are genetically equipped to display lysogenic cycles (temperate versus non-temperate phages). We address in particular the use or overuse of what can be a somewhat equivocal phrase, 'Lytic or lysogenic', especially when employed as a means of distinguishing among phages types. We suggest that the implied dichotomy is inconsistent with both modern as well as historical understanding of phage biology. We consider, therefore, less ambiguous terminology for distinguishing between 'Lytic' versus 'Lysogenic' phage types. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Phages of lactic acid bacteria: The role of genetics in understanding phage-host interactions and their co-evolutionary processes

International Nuclear Information System (INIS)

Mahony, Jennifer; Ainsworth, Stuart; Stockdale, Stephen; Sinderen, Douwe van

2012-01-01

Dairy fermentations are among the oldest food processing applications, aimed at preservation and shelf-life extension through the use of lactic acid bacteria (LAB) starter cultures, in particular strains of Lactococcus lactis, Streptococcus thermophilus, Lactobacillus spp. and Leuconostoc spp. Traditionally this was performed by continuous passaging of undefined cultures from a finished fermentation to initiate the next fermentation. More recently, consumer demands on consistent and desired flavours and textures of dairy products have led to a more defined approach to such processes. Dairy (starter) companies have responded to the need to define the nature and complexity of the starter culture mixes, and dairy fermentations are now frequently based on defined starter cultures of low complexity, where each starter component imparts specific technological properties that are desirable to the product. Both mixed and defined starter culture approaches create the perfect environment for the proliferation of (bacterio)phages capable of infecting these LAB. The repeated use of the same starter cultures in a single plant, coupled to the drive towards higher and consistent production levels, increases the risk and negative impact of phage infection. In this review we will discuss recent advances in tracking the adaptation of phages to the dairy industry, the advances in understanding LAB phage-host interactions, including evolutionary and genomic aspects.

Phages of lactic acid bacteria: The role of genetics in understanding phage-host interactions and their co-evolutionary processes

Energy Technology Data Exchange (ETDEWEB)

Mahony, Jennifer, E-mail: j.mahony@ucc.ie [Department of Microbiology, University College Cork, Western Road, Cork (Ireland); Ainsworth, Stuart; Stockdale, Stephen [Department of Microbiology, University College Cork, Western Road, Cork (Ireland); Sinderen, Douwe van, E-mail: d.vansinderen@ucc.ie [Department of Microbiology, University College Cork, Western Road, Cork (Ireland); Alimentary Pharmabiotic Centre, Biosciences Institute, University College Cork, Western Road, Cork (Ireland)

2012-12-20

Dairy fermentations are among the oldest food processing applications, aimed at preservation and shelf-life extension through the use of lactic acid bacteria (LAB) starter cultures, in particular strains of Lactococcus lactis, Streptococcus thermophilus, Lactobacillus spp. and Leuconostoc spp. Traditionally this was performed by continuous passaging of undefined cultures from a finished fermentation to initiate the next fermentation. More recently, consumer demands on consistent and desired flavours and textures of dairy products have led to a more defined approach to such processes. Dairy (starter) companies have responded to the need to define the nature and complexity of the starter culture mixes, and dairy fermentations are now frequently based on defined starter cultures of low complexity, where each starter component imparts specific technological properties that are desirable to the product. Both mixed and defined starter culture approaches create the perfect environment for the proliferation of (bacterio)phages capable of infecting these LAB. The repeated use of the same starter cultures in a single plant, coupled to the drive towards higher and consistent production levels, increases the risk and negative impact of phage infection. In this review we will discuss recent advances in tracking the adaptation of phages to the dairy industry, the advances in understanding LAB phage-host interactions, including evolutionary and genomic aspects.

Plasmids and packaging cell lines for use in phage display

Science.gov (United States)

Bradbury, Andrew M.

2012-07-24

The invention relates to a novel phagemid display system for packaging phagemid DNA into phagemid particles which completely avoids the use of helper phage. The system of the invention incorporates the use of bacterial packaging cell lines which have been transformed with helper plasmids containing all required phage proteins but not the packaging signals. The absence of packaging signals in these helper plasmids prevents their DNA from being packaged in the bacterial cell, which provides a number of significant advantages over the use of both standard and modified helper phage. Packaged phagemids expressing a protein or peptide of interest, in fusion with a phage coat protein such as g3p, are generated simply by transfecting phagemid into the packaging cell line.

Genomic analysis of WCP30 Phage of Weissella cibaria for Dairy Fermented Foods.

Science.gov (United States)

Lee, Young-Duck; Park, Jong-Hyun

2017-01-01

In this study, we report the morphogenetic analysis and genome sequence of a new WCP30 phage of Weissella cibaria , isolated from a fermented food. Based on its morphology, as observed by transmission electron microscopy, WCP30 phage belongs to the family Siphoviridae . Genomic analysis of WCP30 phage showed that it had a 33,697-bp double-stranded DNA genome with 41.2% G+C content. Bioinformatics analysis of the genome revealed 35 open reading frames. A BLASTN search showed that WCP30 phage had low sequence similarity compared to other phages infecting lactic acid bacteria. This is the first report of the morphological features and complete genome sequence of WCP30 phage, which may be useful for controlling the fermentation of dairy foods.

Oral T4-like phage cocktail application to healthy adult volunteers from Bangladesh

International Nuclear Information System (INIS)

Sarker, Shafiqul Alam; McCallin, Shawna; Barretto, Caroline; Berger, Bernard; Pittet, Anne-Cécile; Sultana, Shamima; Krause, Lutz; Huq, Sayeda; Bibiloni, Rodrigo; Bruttin, Anne; Reuteler, Gloria; Brüssow, Harald

2012-01-01

The genomic diversity of 99 T4-like coliphages was investigated by sequencing an equimolar mixture with Illumina technology and screening them against different databases for horizontal gene transfer and undesired genes. A 9-phage cocktail was given to 15 healthy adults from Bangladesh at a dose of 3×10 9 and 3×10 7 plaque-forming units and placebo respectively. Phages were detected in 64% of the stool samples when subjects were treated with higher titer phage, compared to 30% and 28% with lower-titer phage and placebo, respectively. No Escherichia coli was present in initial stool samples, and no amplification of phage was observed. One percent of the administered oral phage was recovered from the feces. No adverse events were observed by self-report, clinical examination, or from laboratory tests for liver, kidney, and hematology function. No impact of oral phage was seen on the fecal microbiota composition with respect to bacterial 16S rRNA from stool.

Oral T4-like phage cocktail application to healthy adult volunteers from Bangladesh

Energy Technology Data Exchange (ETDEWEB)

Sarker, Shafiqul Alam, E-mail: sasarker@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); McCallin, Shawna; Barretto, Caroline [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Berger, Bernard, E-mail: bernard.berger@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Pittet, Anne-Cecile [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Sultana, Shamima, E-mail: shamima@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Krause, Lutz, E-mail: ltz.krause@gmail.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Huq, Sayeda, E-mail: sayeeda@mail.icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Bibiloni, Rodrigo, E-mail: Rodrigo.Bibiloni@agresearch.co.nz [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Bruttin, Anne, E-mail: anne.bruttin@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Reuteler, Gloria, E-mail: gloria.reuteler@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Bruessow, Harald, E-mail: harald.bruessow@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland)

2012-12-20

The genomic diversity of 99 T4-like coliphages was investigated by sequencing an equimolar mixture with Illumina technology and screening them against different databases for horizontal gene transfer and undesired genes. A 9-phage cocktail was given to 15 healthy adults from Bangladesh at a dose of 3 Multiplication-Sign 10{sup 9} and 3 Multiplication-Sign 10{sup 7} plaque-forming units and placebo respectively. Phages were detected in 64% of the stool samples when subjects were treated with higher titer phage, compared to 30% and 28% with lower-titer phage and placebo, respectively. No Escherichia coli was present in initial stool samples, and no amplification of phage was observed. One percent of the administered oral phage was recovered from the feces. No adverse events were observed by self-report, clinical examination, or from laboratory tests for liver, kidney, and hematology function. No impact of oral phage was seen on the fecal microbiota composition with respect to bacterial 16S rRNA from stool.

Filament Substructures and their Interrelation

Science.gov (United States)

Lin, Y.; Martin, S. F.; Engvold, O.

The main structural components of solar filaments, their spines, barbs, and legs at the extreme ends of the spine, are illustrated from recent high-resolution observations. The thread-like structures appear to be present in filaments everywhere and at all times. They are the fundamental elements of solar filaments. The interrelation of the spines, barbs and legs are discussed. From observations, we present a conceptual model of the magnetic field of a filament. We suggest that only a single physical model is needed to explain filaments in a continuous spectrum represented by active region filaments at one end and quiescent filaments at the other end.

Phage morphology recapitulates phylogeny: the comparative genomics of a new group of myoviruses.

Directory of Open Access Journals (Sweden)

André M Comeau

Full Text Available Among dsDNA tailed bacteriophages (Caudovirales, members of the Myoviridae family have the most sophisticated virion design that includes a complex contractile tail structure. The Myoviridae generally have larger genomes than the other phage families. Relatively few "dwarf" myoviruses, those with a genome size of less than 50 kb such as those of the Mu group, have been analyzed in extenso. Here we report on the genome sequencing and morphological characterization of a new group of such phages that infect a diverse range of Proteobacteria, namely Aeromonas salmonicida phage 56, Vibrio cholerae phages 138 and CP-T1, Bdellovibrio phage φ1422, and Pectobacterium carotovorum phage ZF40. This group of dwarf myoviruses shares an identical virion morphology, characterized by usually short contractile tails, and have genome sizes of approximately 45 kb. Although their genome sequences are variable in their lysogeny, replication, and host adaption modules, presumably reflecting differing lifestyles and hosts, their structural and morphogenesis modules have been evolutionarily constrained by their virion morphology. Comparative genomic analysis reveals that these phages, along with related prophage genomes, form a new coherent group within the Myoviridae. The results presented in this communication support the hypothesis that the diversity of phages may be more structured than generally believed and that the innumerable phages in the biosphere all belong to discrete lineages or families.

The genome of the Erwinia amylovora phage PhiEaH1 reveals greater diversity and broadens the applicability of phages for the treatment of fire blight.

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Meczker, Katalin; Dömötör, Dóra; Vass, János; Rákhely, Gábor; Schneider, György; Kovács, Tamás

2014-01-01

The enterobacterium Erwinia amylovora is the causal agent of fire blight. This study presents the analysis of the complete genome of phage PhiEaH1, isolated from the soil surrounding an E. amylovora-infected apple tree in Hungary. Its genome is 218 kb in size, containing 244 ORFs. PhiEaH1 is the second E. amylovora infecting phage from the Siphoviridae family whose complete genome sequence was determined. Beside PhiEaH2, PhiEaH1 is the other active component of Erwiphage, the first bacteriophage-based pesticide on the market against E. amylovora. Comparative genome analysis in this study has revealed that PhiEaH1 not only differs from the 10 formerly sequenced E. amylovora bacteriophages belonging to other phage families, but also from PhiEaH2. Sequencing of more Siphoviridae phage genomes might reveal further diversity, providing opportunities for the development of even more effective biological control agents, phage cocktails against Erwinia fire blight disease of commercial fruit crops.

Characterization of novel virulent broad-host-range phages of Xylella fastidiosa and Xanthomonas.

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Ahern, Stephen J; Das, Mayukh; Bhowmick, Tushar Suvra; Young, Ry; Gonzalez, Carlos F

2014-01-01

The xylem-limited bacterium Xylella fastidiosa is the causal agent of several plant diseases, most notably Pierce's disease of grape and citrus variegated chlorosis. We report the isolation and characterization of the first virulent phages for X. fastidiosa, siphophages Sano and Salvo and podophages Prado and Paz, with a host range that includes Xanthomonas spp. Phages propagated on homologous hosts had observed adsorption rate constants of ~4 × 10(-12) ml cell(-1) min(-1) for X. fastidiosa strain Temecula 1 and ~5 × 10(-10) to 7 × 10(-10) ml cell(-1) min(-1) for Xanthomonas strain EC-12. Sano and Salvo exhibit >80% nucleotide identity to each other in aligned regions and are syntenic to phage BcepNazgul. We propose that phage BcepNazgul is the founding member of a novel phage type, to which Sano and Salvo belong. The lysis genes of the Nazgul-like phage type include a gene that encodes an outer membrane lipoprotein endolysin and also spanin gene families that provide insight into the evolution of the lysis pathway for phages of Gram-negative hosts. Prado and Paz, although exhibiting no significant DNA homology to each other, are new members of the phiKMV-like phage type, based on the position of the single-subunit RNA polymerase gene. The four phages are type IV pilus dependent for infection of both X. fastidiosa and Xanthomonas. The phages may be useful as agents for an effective and environmentally responsible strategy for the control of diseases caused by X. fastidiosa.

The True Story and Advantages of RNA Phage Capsids as Nanotools.

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Pumpens, Paul; Renhofa, Regina; Dishlers, Andris; Kozlovska, Tatjana; Ose, Velta; Pushko, Peter; Tars, Kaspars; Grens, Elmars; Bachmann, Martin F

2016-01-01

RNA phages are often used as prototypes for modern recombinant virus-like particle (VLP) technologies. Icosahedral RNA phage VLPs can be formed from coat proteins (CPs) and are efficiently produced in bacteria and yeast. Both genetic fusion and chemical coupling have been successfully used for the production of numerous chimeras based on RNA phage VLPs. In this review, we describe advances in RNA phage VLP technology along with the history of the Leviviridae family, including its taxonomical organization, genomic structure, and important role in the development of molecular biology. Comparative 3D structures of different RNA phage VLPs are used to explain the level of VLP tolerance to foreign elements displayed on VLP surfaces. We also summarize data that demonstrate the ability of CPs to tolerate different organic (peptides, oligonucleotides, and carbohydrates) and inorganic (metal ions) compounds either chemically coupled or noncovalently added to the outer and/or inner surfaces of VLPs. Finally, we present lists of nanotechnological RNA phage VLP applications, such as experimental vaccines constructed by genetic fusion and chemical coupling methodologies, nanocontainers for targeted drug delivery, and bioimaging tools. © 2016 S. Karger AG, Basel.

Recovery of phage lambda from ultraviolet damage

International Nuclear Information System (INIS)

Devoret, R.; Blanco, M.; George, J.; Radman, M.

1975-01-01

Recovery of phage lambda from ultraviolet damage can occur, in the dark, through three types of repair processes as defined by microbiological tests: host-cell reactivation, prophage reactivation, and uv reactivation. This paper reviews the properties of the three repair processes, analyzes their dependence on the functioning of bacterial and phage genes, and discusses their relationship. Progress in the understanding of the molecular mechanisms underlying the three repair processes has been relatively slow, particularly for uv reactivation. It has been shown that host-cell reactivation is due to pyrimidine dimer excision and that prophage reactivation is due to genetic recombination (prereplicative). We provide evidence showing that neither of these mechanisms accounts for uv reactivation of phage lambda. Furthermore, uv reactivation differs from the other repair processes in that it is inducible and error-prone. Whether uv-damaged bacterial DNA is subject to a similar repair process is still an open question

Phage based green chemistry for gold ion reduction and gold retrieval.

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Setyawati, Magdiel I; Xie, Jianping; Leong, David T

2014-01-22

The gold mining industry has taken its toll on the environment, triggering the development of more environmentally benign processes to alleviate the waste load release. Here, we demonstrate the use of bacteriophages (phages) for biosorption and bioreduction of gold ions from aqueous solution, which potentially can be applied to remediate gold ions from gold mining waste effluent. Phage has shown a remarkably efficient sorption of gold ions with a maximum gold adsorption capacity of 571 mg gold/g dry weight phage. The product of this phage mediated process is gold nanocrystals with the size of 30-630 nm. Biosorption and bioreduction processes are mediated by the ionic and covalent interaction between gold ions and the reducing groups on the phage protein coat. The strategy offers a simple, ecofriendly and feasible option to recover of gold ions to form readily recoverable products of gold nanoparticles within 24 h.

Metagenomic recovery of phage genomes of uncultured freshwater actinobacteria.

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Ghai, Rohit; Mehrshad, Maliheh; Mizuno, Carolina Megumi; Rodriguez-Valera, Francisco

2017-01-01

Low-GC Actinobacteria are among the most abundant and widespread microbes in freshwaters and have largely resisted all cultivation efforts. Consequently, their phages have remained totally unknown. In this work, we have used deep metagenomic sequencing to assemble eight complete genomes of the first tailed phages that infect freshwater Actinobacteria. Their genomes encode the actinobacterial-specific transcription factor whiB, frequently found in mycobacteriophages and also in phages infecting marine pelagic Actinobacteria. Its presence suggests a common and widespread strategy of modulation of host transcriptional machinery upon infection via this transcriptional switch. We present evidence that some whiB-carrying phages infect the acI lineage of Actinobacteria. At least one of them encodes the ADP-ribosylating component of the widespread bacterial AB toxins family (for example, clostridial toxin). We posit that the presence of this toxin reflects a 'trojan horse' strategy, providing protection at the population level to the abundant host microbes against eukaryotic predators.

A highly specific phage defense system is a conserved feature of the Vibrio cholerae mobilome.

Directory of Open Access Journals (Sweden)

Brendan J O'Hara

2017-06-01

Full Text Available Vibrio cholerae-specific bacteriophages are common features of the microbial community during cholera infection in humans. Phages impose strong selective pressure that favors the expansion of phage-resistant strains over their vulnerable counterparts. The mechanisms allowing virulent V. cholerae strains to defend against the ubiquitous threat of predatory phages have not been established. Here, we show that V. cholerae PLEs (phage-inducible chromosomal island-like elements are widespread genomic islands dedicated to phage defense. Analysis of V. cholerae isolates spanning a 60-year collection period identified five unique PLEs. Remarkably, we found that all PLEs (regardless of geographic or temporal origin respond to infection by a myovirus called ICP1, the most prominent V. cholerae phage found in cholera patient stool samples from Bangladesh. We found that PLE activity reduces phage genome replication and accelerates cell lysis following ICP1 infection, killing infected host cells and preventing the production of progeny phage. PLEs are mobilized by ICP1 infection and can spread to neighboring cells such that protection from phage predation can be horizontally acquired. Our results reveal that PLEs are a persistent feature of the V. cholerae mobilome that are adapted to providing protection from a single predatory phage and advance our understanding of how phages influence pathogen evolution.

A highly specific phage defense system is a conserved feature of the Vibrio cholerae mobilome.

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O'Hara, Brendan J; Barth, Zachary K; McKitterick, Amelia C; Seed, Kimberley D

2017-06-01

Vibrio cholerae-specific bacteriophages are common features of the microbial community during cholera infection in humans. Phages impose strong selective pressure that favors the expansion of phage-resistant strains over their vulnerable counterparts. The mechanisms allowing virulent V. cholerae strains to defend against the ubiquitous threat of predatory phages have not been established. Here, we show that V. cholerae PLEs (phage-inducible chromosomal island-like elements) are widespread genomic islands dedicated to phage defense. Analysis of V. cholerae isolates spanning a 60-year collection period identified five unique PLEs. Remarkably, we found that all PLEs (regardless of geographic or temporal origin) respond to infection by a myovirus called ICP1, the most prominent V. cholerae phage found in cholera patient stool samples from Bangladesh. We found that PLE activity reduces phage genome replication and accelerates cell lysis following ICP1 infection, killing infected host cells and preventing the production of progeny phage. PLEs are mobilized by ICP1 infection and can spread to neighboring cells such that protection from phage predation can be horizontally acquired. Our results reveal that PLEs are a persistent feature of the V. cholerae mobilome that are adapted to providing protection from a single predatory phage and advance our understanding of how phages influence pathogen evolution.

Analysis of whole genome sequencing for the Escherichia coli O157:H7 typing phages.

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Cowley, Lauren A; Beckett, Stephen J; Chase-Topping, Margo; Perry, Neil; Dallman, Tim J; Gally, David L; Jenkins, Claire

2015-04-08

Shiga toxin producing Escherichia coli O157 can cause severe bloody diarrhea and haemolytic uraemic syndrome. Phage typing of E. coli O157 facilitates public health surveillance and outbreak investigations, certain phage types are more likely to occupy specific niches and are associated with specific age groups and disease severity. The aim of this study was to analyse the genome sequences of 16 (fourteen T4 and two T7) E. coli O157 typing phages and to determine the genes responsible for the subtle differences in phage type profiles. The typing phages were sequenced using paired-end Illumina sequencing at The Genome Analysis Centre and the Animal Health and Veterinary Laboratories Agency and bioinformatics programs including Velvet, Brig and Easyfig were used to analyse them. A two-way Euclidian cluster analysis highlighted the associations between groups of phage types and typing phages. The analysis showed that the T7 typing phages (9 and 10) differed by only three genes and that the T4 typing phages formed three distinct groups of similar genomic sequences: Group 1 (1, 8, 11, 12 and 15, 16), Group 2 (3, 6, 7 and 13) and Group 3 (2, 4, 5 and 14). The E. coli O157 phage typing scheme exhibited a significantly modular network linked to the genetic similarity of each group showing that these groups are specialised to infect a subset of phage types. Sequencing the typing phage has enabled us to identify the variable genes within each group and to determine how this corresponds to changes in phage type.

Production of a phage-displayed single chain variable fragment ...

African Journals Online (AJOL)

Purpose: To develop specific single chain variable fragments (scFv) against infectious bursal disease virus (IBDV) via phage display technology. Methods: Purified viruses were initially applied for iterative panning rounds of scFv phage display libraries. The binding ability of the selected scFv antibody fragments against the ...

Bacteriophage-resistant mutants in Yersinia pestis: identification of phage receptors and attenuation for mice.

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Andrey A Filippov

Full Text Available BACKGROUND: Bacteriophages specific for Yersinia pestis are routinely used for plague diagnostics and could be an alternative to antibiotics in case of drug-resistant plague. A major concern of bacteriophage therapy is the emergence of phage-resistant mutants. The use of phage cocktails can overcome this problem but only if the phages exploit different receptors. Some phage-resistant mutants lose virulence and therefore should not complicate bacteriophage therapy. METHODOLOGY/PRINCIPAL FINDINGS: The purpose of this work was to identify Y. pestis phage receptors using site-directed mutagenesis and trans-complementation and to determine potential attenuation of phage-resistant mutants for mice. Six receptors for eight phages were found in different parts of the lipopolysaccharide (LPS inner and outer core. The receptor for R phage was localized beyond the LPS core. Most spontaneous and defined phage-resistant mutants of Y. pestis were attenuated, showing increase in LD₅₀ and time to death. The loss of different LPS core biosynthesis enzymes resulted in the reduction of Y. pestis virulence and there was a correlation between the degree of core truncation and the impact on virulence. The yrbH and waaA mutants completely lost their virulence. CONCLUSIONS/SIGNIFICANCE: We identified Y. pestis receptors for eight bacteriophages. Nine phages together use at least seven different Y. pestis receptors that makes some of them promising for formulation of plague therapeutic cocktails. Most phage-resistant Y. pestis mutants become attenuated and thus should not pose a serious problem for bacteriophage therapy of plague. LPS is a critical virulence factor of Y. pestis.

Treatment of in vitro enterohemorrhagic Escherichia coli infection using phage and probiotics.

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Dini, C; Bolla, P A; de Urraza, P J

2016-07-01

To assay the combination of phage and probiotics against EHEC in vitro on infected Hep-2 cells. Phage and probiotics treatments on EHEC O157:H7-infected Hep-2 cells were assayed individually or combined. The effect of freeze-drying on phage and probiotic antimicrobial activity was also studied. While treatment with phage alone increased cell detachment caused by EHEC infection, the treatments with MM alone or in combination with phage proved to effectively diminish cell damage caused by EHEC infection. Combined treatment showed a decrease in apoptotic cell count of 57·3% and a reduction in EHEC adhesion to cell monolayer of 1·2 log CFU. The simultaneous use of phage and probiotics showed no antagonistic effect, and freeze-drying did not affect their antipathogenic activity. The combination of phage and probiotics has great potential for reducing the number of pathogens adhered to epithelial cells during EHEC O157:H7 infection and attenuating the cytotoxic effect derived from it. Further in vivo assays are needed for assessing the actual effectiveness of the treatment. This study presents a freeze-dried formulation of phage and probiotics capable of controlling EHEC infections and reducing epithelial cell damage in vitro. © 2016 The Society for Applied Microbiology.

Survey on the phage resistance mechanisms displayed by a dairy Lactobacillus helveticus strain.

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Zago, Miriam; Orrù, Luigi; Rossetti, Lia; Lamontanara, Antonella; Fornasari, Maria Emanuela; Bonvini, Barbara; Meucci, Aurora; Carminati, Domenico; Cattivelli, Luigi; Giraffa, Giorgio

2017-09-01

In this study the presence and functionality of phage defence mechanisms in Lactobacillus helveticus ATCC 10386, a strain of dairy origin which is sensitive to ΦLh56, were investigated. After exposure of ATCC 10386 to ΦLh56, the whole-genome sequences of ATCC 10386 and of a phage-resistant derivative (LhM3) were compared. LhM3 showed deletions in the S-layer protein and a higher expression of the genes involved in the restriction/modification (R/M) system. Genetic data were substantiated by measurements of bacteriophage adsorption rates, efficiency of plaquing, cell wall protein size and by gene expression analysis. In LhM3 two phage resistance mechanisms, the inhibition of phage adsorption and the upregulation of Type I R/M genes, take place and explain its resistance to ΦLh56. Although present in both ATCC 10386 and LhM3 genomes, the CRISPR machinery did not seem to play a role in the phage resistance of LhM3. Overall, the natural selection of phage resistant strains resulted successful in detecting variants carrying multiple phage defence mechanisms in L. helveticus. The concurrent presence of multiple phage-resistance systems should provide starter strains with increased fitness and robustness in dairy ecosystems. Copyright © 2017 Elsevier Ltd. All rights reserved.

Novel Variants of Streptococcus thermophilus Bacteriophages Are Indicative of Genetic Recombination among Phages from Different Bacterial Species

DEFF Research Database (Denmark)

Szymczak, Paula; Janzen, Thomas; Neves, Ana Rute

2017-01-01

lactis P335 phages. Phage CHPC1151 was closely related to the atypical S. thermophilus phage 5093, homologous with a nondairy streptococcal prophage. By testing adsorption of the related streptococcal and lactococcal phages to the surface of S. thermophilus and L. lactis strains, we revealed....... thermophilus phages from the Chr. Hansen A/S collection, using PCR specific for the cos- or pac-type phages, as well as for the V2 antireceptor region. Three phages did not produce positive results with the assays. Analysis of phage morphologies indicated that two of these phages, CHPC577 and CHPC926, had...... the possibility of cross-interactions. Our data indicated that the use of S. thermophilus together with L. lactis, extensively applied for dairy fermentations, triggered the recombination between phages infecting different bacterial species. A notable diversity among S. thermophilus phage populations requires...

SYMPATHETIC SOLAR FILAMENT ERUPTIONS

Energy Technology Data Exchange (ETDEWEB)

Wang, Rui; Liu, Ying D.; Zimovets, Ivan; Hu, Huidong; Yang, Zhongwei [State Key Laboratory of Space Weather, National Space Science Center, Chinese Academy of Sciences, Beijing 100190 (China); Dai, Xinghua, E-mail: liuxying@spaceweather.ac.cn [Key Laboratory of Solar Activity, National Astronomical Observatories, Chinese Academy of Sciences, Beijing 100012 (China)

2016-08-10

The 2015 March 15 coronal mass ejection as one of the two that together drove the largest geomagnetic storm of solar cycle 24 so far was associated with sympathetic filament eruptions. We investigate the relations between the different filaments involved in the eruption. A surge-like small-scale filament motion is confirmed as the trigger that initiated the erupting filament with multi-wavelength observations and using a forced magnetic field extrapolation method. When the erupting filament moved to an open magnetic field region, it experienced an obvious acceleration process and was accompanied by a C-class flare and the rise of another larger filament that eventually failed to erupt. We measure the decay index of the background magnetic field, which presents a critical height of 118 Mm. Combining with a potential field source surface extrapolation method, we analyze the distributions of the large-scale magnetic field, which indicates that the open magnetic field region may provide a favorable condition for F2 rapid acceleration and have some relation with the largest solar storm. The comparison between the successful and failed filament eruptions suggests that the confining magnetic field plays an important role in the preconditions for an eruption.

Modern filaments for composite materials

International Nuclear Information System (INIS)

Krivelli-Viskonti, I.

1982-01-01

Analysis of modern state and ways to improve properties of different filaments for the forecast of the filament application in composite materials has been conducted. In the near future as before the greatest attention will be paid to fibre glass, as this material is widely used in the reinforcing of organic matrices. Carbon and kevlar filaments are the most prospective ones. For the service at medium, high or superhigh temperatures selection of matrix material is more significant than selection of filament. Organic matrices can not be used at temperatures > 250 deg C: this is already the range of metal matrix application. Though at temperatures above room one many filaments can be used, boron filaments and metal wire are the only reinforcing materials, inspite of the fact that carbon filaments are successfully used for metal matrix reinforcing. At very high temperatures only carbon filaments or silicon carbide ones can be used, but their cost is very high and besides economical problems there are many difficulties of technical character

Understanding the enormous diversity of bacteriophages: the tailed phages that infect the bacterial family Enterobacteriaceae

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Grose, Julianne H.; Casjens, Sherwood R.

2014-01-01

Bacteriophages are the predominant biological entity on the planet. The recent explosion of sequence information has made estimates of their diversity possible. We describe the genomic comparison of 337 fully sequenced tailed phages isolated on 18 genera and 31 species of bacteria in the Enterobacteriaceae. These phages were largely unambiguously grouped into 56 diverse clusters (32 lytic and 24 temperate) that have syntenic similarity over >50% of the genomes within each cluster, but substantially less sequence similarity between clusters. Most clusters naturally break into sets of more closely related subclusters, 78% of which are correlated with their host genera. The largest groups of related phages are superclusters united by genome synteny to lambda (81 phages) and T7 (51 phages). This study forms a robust framework for understanding diversity and evolutionary relationships of existing tailed phages, for relating newly discovered phages and for determining host/phage relationships. PMID:25240328

Phage Therapy Approaches to Reducing Pathogen Persistence and Transmission in Animal Production Environments: Opportunities and Challenges.

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Colavecchio, Anna; Goodridge, Lawrence D

2017-06-01

The era of genomics has allowed for characterization of phages for use as antimicrobials to treat animal infections with a level of precision never before realized. As more research in phage therapy has been conducted, several advantages of phage therapy have been realized, including the ubiquitous nature, specificity, prevalence in the biosphere, and low inherent toxicity of phages, which makes them a safe and sustainable technology for control of animal diseases. These unique qualities of phages have led to several opportunities with respect to emerging trends in infectious disease treatment. However, the opportunities are tempered by several challenges to the successful implementation of phage therapy, such as the fact that an individual phage can only infect one or a few bacterial strains, meaning that large numbers of different phages will likely be needed to treat infections caused by multiple species of bacteria. In addition, phages are only effective if enough of them can reach the site of bacterial colonization, but clearance by the immune system upon introduction to the animal is a reality that must be overcome. Finally, bacterial resistance to the phages may develop, resulting in treatment failure. Even a successful phage infection and lysis of its host has consequences, because large amounts of endotoxin are released upon lysis of Gram-negative bacteria, which can lead to local and systemic complications. Overcoming these challenges will require careful design and development of phage cocktails, including comprehensive characterization of phage host range and assessment of immunological risks associated with phage treatment.

Induced alignment and measurement of dipolar couplings of an SH2 domain through direct binding with filamentous phage

International Nuclear Information System (INIS)

Dahlke Ojennus, Deanna; Mitton-Fry, Rachel M.; Wuttke, Deborah S.

1999-01-01

Large residual 15 N- 1 H dipolar couplings have been measured in a Src homology II domain aligned at Pf1 bacteriophage concentrations an order of magnitude lower than used for induction of a similar degree of alignment of nucleic acids and highly acidic proteins. An increase in 1 H and 15 N protein linewidths and a decrease in T 2 and T 1 ρ relaxation time constants implicates a binding interaction between the protein and phage as the mechanism of alignment. However, the associated increased linewidth does not preclude the accurate measurement of large dipolar couplings in the aligned protein. A good correlation is observed between measured dipolar couplings and predicted values based on the high resolution NMR structure of the SH2 domain. The observation of binding-induced protein alignment promises to broaden the scope of alignment techniques by extending their applicability to proteins that are able to interact weakly with the alignment medium

Epifluorescence and atomic force microscopy: Two innovative applications for studying phage-host interactions in Lactobacillus helveticus.

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Zago, Miriam; Scaltriti, Erika; Fornasari, Maria Emanuela; Rivetti, Claudio; Grolli, Stefano; Giraffa, Giorgio; Ramoni, Roberto; Carminati, Domenico

2012-01-01

Bacteriophages attacking lactic acid bacteria (LAB) still represent a crucial problem in industrial dairy fermentations. The consequences of a phage infection against LAB can lead to fermentation delay, alteration of the product quality and, in most severe cases, the product loss. Phage particles enumeration and phage-host interactions are normally evaluated by conventional plaque count assays, but, in many cases, these methods can be unsuccessful. Bacteriophages of Lactobacillus helveticus, a LAB species widely used as dairy starter or probiotic cultures, are often unable to form lysis plaques, thus impairing their enumeration by plate assay. In this study, we used epifluorescence microscopy to enumerate L. helveticus phage particles from phage-infected cultures and Atomic Force Microscopy (AFM) to visualize both phages and bacteria during the different stages of the lytic cycle. Preliminary, we tested the sensitivity of phage counting by epifluorescence microscopy. To this end, phage particles of ΦAQ113, a lytic phage of L. helveticus isolated from a whey starter culture, were stained by SYBR Green I and enumerated by epifluorescence microscopy. Values obtained by the microscopic method were 10 times higher than plate counts, with a lowest sensitivity limit of ≥6log phage/ml. The interaction of phage ΦAQ113 with its host cell L. helveticus Lh1405 was imaged by AFM after 0, 2 and 5h from phage-host adsorption. The lytic cycle was followed by epifluorescence microscopy counting and the concomitant cell wall changes were visualized by AFM imaging. Our results showed that these two methods can be combined for a reliable phage enumeration and for studying phage and host morphology during infection processes, thus giving a complete overview of phage-host interactions in L. helveticus strains involved in dairy productions. Copyright © 2011 Elsevier B.V. All rights reserved.

Heterogeneity in Induction Level, Infection Ability, and Morphology of Shiga Toxin-Encoding Phages (Stx Phages) from Dairy and Human Shiga Toxin-Producing Escherichia coli O26:H11 Isolates

Science.gov (United States)

Bonanno, Ludivine; Petit, Marie-Agnès; Loukiadis, Estelle; Michel, Valérie

2016-01-01

Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are foodborne pathogens responsible for diarrhea and hemolytic-uremic syndrome (HUS). Shiga toxin, the main STEC virulence factor, is encoded by the stx gene located in the genome of a bacteriophage inserted into the bacterial chromosome. The O26:H11 serotype is considered to be the second-most-significant HUS-causing serotype worldwide after O157:H7. STEC O26:H11 bacteria and their stx-negative counterparts have been detected in dairy products. They may convert from the one form to the other by loss or acquisition of Stx phages, potentially confounding food microbiological diagnostic methods based on stx gene detection. Here we investigated the diversity and mobility of Stx phages from human and dairy STEC O26:H11 strains. Evaluation of their rate of in vitro induction, occurring either spontaneously or in the presence of mitomycin C, showed that the Stx2 phages were more inducible overall than Stx1 phages. However, no correlation was found between the Stx phage levels produced and the origin of the strains tested or the phage insertion sites. Morphological analysis by electron microscopy showed that Stx phages from STEC O26:H11 displayed various shapes that were unrelated to Stx1 or Stx2 types. Finally, the levels of sensitivity of stx-negative E. coli O26:H11 to six Stx phages differed among the 17 strains tested and our attempts to convert them into STEC were unsuccessful, indicating that their lysogenization was a rare event. PMID:26826235

Phage Conversion for β-Lactam Antibiotic Resistance of Staphylococcus aureus from Foods.

Science.gov (United States)

Lee, Young-Duck; Park, Jong-Hyun

2016-02-01

Temperate phages have been suggested to carry virulence factors and other lysogenic conversion genes that play important roles in pathogenicity. In this study, phage TEM123 in wild-type Staphylococcus aureus from food sources was analyzed with respect to its morphology, genome sequence, and antibiotic resistance conversion ability. Phage TEM123 from a mitomycin C-induced lysate of S. aureus was isolated from foods. Morphological analysis under a transmission electron microscope revealed that it belonged to the family Siphoviridae. The genome of phage TEM123 consisted of a double-stranded DNA of 43,786 bp with a G+C content of 34.06%. A bioinformatics analysis of the phage genome identified 43 putative open reading frames (ORFs). ORF1 encoded a protein that was nearly identical to the metallo-β-lactamase enzymes that degrade β-lactam antibiotics. After transduction to S. aureus with phage TEM123, the metallo-β-lactamase gene was confirmed in the transductant by PCR and sequencing analyses. In a β-lactam antibiotic susceptibility test, the transductant was more highly resistant to β-lactam antibiotics than S. aureus S133. Phage TEM123 might play a role in the transfer of β-lactam antibiotic resistance determinants in S. aureus. Therefore, we suggest that the prophage of S. aureus with its exotoxin is a risk factor for food safety in the food chain through lateral gene transfer.

Redundancy and cooperativity in the mechanics of compositely crosslinked filamentous networks.

Directory of Open Access Journals (Sweden)

Moumita Das

Full Text Available The cytoskeleton of living cells contains many types of crosslinkers. Some crosslinkers allow energy-free rotations between filaments and others do not. The mechanical interplay between these different crosslinkers is an open issue in cytoskeletal mechanics. Therefore, we develop a theoretical framework based on rigidity percolation to study a generic filamentous system containing both stretching and bond-bending forces to address this issue. The framework involves both analytical calculations via effective medium theory and numerical simulations on a percolating triangular lattice with very good agreement between both. We find that the introduction of angle-constraining crosslinkers to a semiflexible filamentous network with freely rotating crosslinks can cooperatively lower the onset of rigidity to the connectivity percolation threshold-a result argued for years but never before obtained via effective medium theory. This allows the system to ultimately attain rigidity at the lowest concentration of material possible. We further demonstrate that introducing angle-constraining crosslinks results in mechanical behaviour similar to just freely rotating crosslinked semflexible filaments, indicating redundancy and universality. Our results also impact upon collagen and fibrin networks in biological and bio-engineered tissues.

Phage-inducible chromosomal islands are ubiquitous within the bacterial universe.

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Fillol-Salom, Alfred; Martínez-Rubio, Roser; Abdulrahman, Rezheen F; Chen, John; Davies, Robert; Penadés, José R

2018-06-06

Phage-inducible chromosomal islands (PICIs) are a recently discovered family of pathogenicity islands that contribute substantively to horizontal gene transfer, host adaptation and virulence in Gram-positive cocci. Here we report that similar elements also occur widely in Gram-negative bacteria. As with the PICIs from Gram-positive cocci, their uniqueness is defined by a constellation of features: unique and specific attachment sites, exclusive PICI genes, a phage-dependent mechanism of induction, conserved replication origin organization, convergent mechanisms of phage interference, and specific packaging of PICI DNA into phage-like infectious particles, resulting in very high transfer frequencies. We suggest that the PICIs represent two or more distinct lineages, have spread widely throughout the bacterial world, and have diverged much more slowly than their host organisms or their prophage cousins. Overall, these findings represent the discovery of a universal class of mobile genetic elements.

Longitudinal monitoring of Listeria monocytogenes and Listeria phages in seafood processing environments in Thailand.

Science.gov (United States)

Vongkamjan, Kitiya; Benjakul, Soottawat; Kim Vu, Hue Thi; Vuddhakul, Varaporn

2017-09-01

Listeria monocytogenes is a foodborne pathogen commonly found in environments of seafood processing, thus presenting a challenge for eradication from seafood processing facilities. Monitoring the prevalence and subtype diversity of L. monocytogenes together with phages that are specific to Listeria spp. ("Listeria phages") will provide knowledge on the bacteria-phage ecology in food processing plants. In this work, a total of 595 samples were collected from raw material, finished seafood products and environmental samples from different sites of a seafood processing plant during 17 sampling visits in 1.5 years of study. L. monocytogenes and Listeria spp. (non-monocytogenes) were found in 22 (3.7%) and 43 (7.2%) samples, respectively, whereas 29 Listeria phages were isolated from 9 (1.5%) phage-positive samples. DNA fingerprint analysis of L. monocytogenes isolates revealed 11 Random Amplified Polymorphic DNA (RAPD) profiles, with two subtypes were frequently observed over time. Our data reveal a presence of Listeria phages within the same seafood processing environments where a diverse set of L. monocytogenes subtypes was also found. Although serotype 4b was observed at lower frequency, data indicate that isolates from this seafood processing plant belonged to both epidemiologically important serotypes 1/2a and 4b, which may suggest a potential public health risk. Phages (all showed a unique genome size of 65 ± 2 kb) were classified into 9 host range groups, representing both broad- and narrow-host range. While most L. monocytogenes isolates from this facility were susceptible to phages, five isolates showed resistance to 12-20 phages. Variations in phage host range among Listeria phages isolated from food processing plant may affect a presence of a diverse set of L. monocytogenes isolates derived from the same processing environment in Thailand. Copyright © 2017 Elsevier Ltd. All rights reserved.

Lethal effects of 32P decay on transfecting activity of Bacillus subtillis phage phie DNA

International Nuclear Information System (INIS)

Loveday, K.S.

1979-01-01

Disintegration of 32 P present in the DNA of Bacillus subtilis phage phie (a phage containing double-strand DNA) results in the loss of viability of intact phage as well as transfecting activity of isolated DNA. Only 1/12 of the 32 P disintegrations per phage DNA equivalent inactivities the intact phage while nearly every disintegration inactivates the transfecting DNA. This result provides evidence for a single-strand intermediate in the transfection of B. subtilis by phie DNA

Exploring the mycobacteriophage metaproteome: phage genomics as an educational platform.

Directory of Open Access Journals (Sweden)

Graham F Hatfull

2006-06-01

Full Text Available Bacteriophages are the most abundant forms of life in the biosphere and carry genomes characterized by high genetic diversity and mosaic architectures. The complete sequences of 30 mycobacteriophage genomes show them collectively to encode 101 tRNAs, three tmRNAs, and 3,357 proteins belonging to 1,536 "phamilies" of related sequences, and a statistical analysis predicts that these represent approximately 50% of the total number of phamilies in the mycobacteriophage population. These phamilies contain 2.19 proteins on average; more than half (774 of them contain just a single protein sequence. Only six phamilies have representatives in more than half of the 30 genomes, and only three-encoding tape-measure proteins, lysins, and minor tail proteins-are present in all 30 phages, although these phamilies are themselves highly modular, such that no single amino acid sequence element is present in all 30 mycobacteriophage genomes. Of the 1,536 phamilies, only 230 (15% have amino acid sequence similarity to previously reported proteins, reflecting the enormous genetic diversity of the entire phage population. The abundance and diversity of phages, the simplicity of phage isolation, and the relatively small size of phage genomes support bacteriophage isolation and comparative genomic analysis as a highly suitable platform for discovery-based education.

Sequential effect of phages and cold nitrogen plasma against Escherichia coli O157:H7 biofilms on different vegetables.

Science.gov (United States)

Cui, Haiying; Bai, Mei; Yuan, Lu; Surendhiran, Duraiarasan; Lin, Lin

2018-03-02

Escherichia coli O157:H7 (E. coli O157:H7) is one of the most common pathogens in fresh vegetables and fruits, and most of the diseases produced by E. coli O157:H7 are associated with biofilms. Cold nitrogen plasma (CNP) is a cold sterilization technique which has no residue. However to completely eliminate the biofilm on the surface of vegetables the processing power and time of CNP have to be enhanced, which will impact on the quality of fruits and vegetables. Thus the sequential treatment of CNP and phage techniques was engineered in this study. Compared to treatment performed separately, sequential treatment not only had more mild treatment conditions as 400W CNP treatment for 2min and 5% phage treatment for 30min, but also exhibited more remarkable effect on eradicating E. coli O157:H7 biofilms in vitro and on vegetables. The population of E. coli O157:H7 was approximately reduced by 2logCFU/cm 2 after individual treatment of 5% phages for 30min or 500W CNP for 3min. While the sequential treatment of CNP (400W, 2min) and phages (5%, 30min) reduced the E. coli O157:H7 viable count in biofilm by 5.71logCFU/cm 2 . Therefore, the sequential treatment holds a great promise to improve the current treatment systems of bacterial contamination on different vegetable surfaces. Copyright © 2018 Elsevier B.V. All rights reserved.

Comparative Omics and Trait Analyses of Marine Pseudoalteromonas Phages Advance the Phage OTU Concept

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Melissa B. Duhaime

2017-07-01

Full Text Available Viruses influence the ecology and evolutionary trajectory of microbial communities. Yet our understanding of their roles in ecosystems is limited by the paucity of model systems available for hypothesis generation and testing. Further, virology is limited by the lack of a broadly accepted conceptual framework to classify viral diversity into evolutionary and ecologically cohesive units. Here, we introduce genomes, structural proteomes, and quantitative host range data for eight Pseudoalteromonas phages isolated from Helgoland (North Sea, Germany and use these data to advance a genome-based viral operational taxonomic unit (OTU definition. These viruses represent five new genera and inform 498 unaffiliated or unannotated protein clusters (PCs from global virus metagenomes. In a comparison of previously sequenced Pseudoalteromonas phage isolates (n = 7 and predicted prophages (n = 31, the eight phages are unique. They share a genus with only one other isolate, Pseudoalteromonas podophage RIO-1 (East Sea, South Korea and two Pseudoalteromonas prophages. Mass-spectrometry of purified viral particles identified 12–20 structural proteins per phage. When combined with 3-D structural predictions, these data led to the functional characterization of five previously unidentified major capsid proteins. Protein functional predictions revealed mechanisms for hijacking host metabolism and resources. Further, they uncovered a hybrid sipho-myovirus that encodes genes for Mu-like infection rarely described in ocean systems. Finally, we used these data to evaluate a recently introduced definition for virus populations that requires members of the same population to have >95% average nucleotide identity across at least 80% of their genes. Using physiological traits and genomics, we proposed a conceptual model for a viral OTU definition that captures evolutionarily cohesive and ecologically distinct units. In this trait-based framework, sensitive hosts are

Phage FR38 Treatment on Sprague Dawley Rat Inferred from Blood Parameters and Organ Systems

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DEWI SARTIKA

2012-09-01

Full Text Available The ability of phage FR38 to lysis indigenous Salmonella P38 from feces of diarrheal patient has been studied. However, effects of phage FR38 on organ system were not revealed as yet. This study was conducted to observe the effect of phage FR38 on blood chemistry, kidney functions, and liver functions. Twelve Sprague-Dawley rats were used as a model for this study that were divided into two groups; (i control and (ii treated group with phage FR38. For treated phage group, each rat was administered by 5 ml/kg bw of 1.59•107 pfu/ml of phage intragastric. The blood parameters were analysed on day 16. The results revealed that body and organs weight, erythrocyte, hematocrit, hemoglobin, leukocyte, total protein, creatinine, SGOT, and SGPT of phage treatment rats were not significantly different with the control rats on day 16 (P > 0.05. Therefore, this study showed was no effect of phage FR38 on body weight, blood chemistry, kidney and liver functions of the rat (P > 0.05.

Phage FR38 Treatment on Sprague Dawley Rat Inferred from Blood Parameters and Organ Systems

Directory of Open Access Journals (Sweden)

DEWI SARTIKA

2012-09-01

Full Text Available The ability of phage FR38 to lysis indigenous Salmonella P38 from feces of diarrheal patient has been studied. However, effects of phage FR38 on organ system were not revealed as yet. This study was conducted to observe the effect of phage FR38 on blood chemistry, kidney functions, and liver functions. Twelve Sprague-Dawley rats were used as a model for this study that were divided into two groups; (i control and (ii treated group with phage FR38. For treated phage group, each rat was administered by 5 ml/kg bw of 1.59-107 pfu/ml of phage intragastric. The blood parameters were analysed on day 16. The results revealed that body and organs weight, erythrocyte, hematocrit, hemoglobin, leukocyte, total protein, creatinine, SGOT, and SGPT of phage treatment rats were not significantly different with the control rats on day 16 (P > 0.05. Therefore, this study showed was no effect of phage FR38 on body weight, blood chemistry, kidney and liver functions of the rat (P > 0.05.

Bioinformatic analysis of phage AB3, a phiKMV-like virus infecting Acinetobacter baumannii.

Science.gov (United States)

Zhang, J; Liu, X; Li, X-J

2015-01-16

The phages of Acinetobacter baumannii has drawn increasing attention because of the multi-drug resistance of A. baumanni. The aim of this study was to sequence Acinetobacter baumannii phage AB3 and conduct bioinformatic analysis to lay a foundation for genome remodeling and phage therapy. We isolated and sequenced A. baumannii phage AB3 and attempted to annotate and analyze its genome. The results showed that the genome is a double-stranded DNA with a total length of 31,185 base pairs (bp) and 97 open reading frames greater than 100 bp. The genome includes 28 predicted genes, of which 24 are homologous to phage AB1. The entire coding sequence is located on the negative strand, representing 90.8% of the total length. The G+C mol% was 39.18%, without areas of high G+C content over 200 bp in length. No GC island, tRNA gene, or repeated sequence was identified. Gene lengths were 120-3099 bp, with an average of 1011 bp. Six genes were found to be greater than 2000 bp in length. Genomic alignment and phylogenetic analysis of the RNA polymerase gene showed that similar to phage AB1, phage AB3 is a phiKMV-like virus in the T7 phage family.

Solar Filament Extraction and Characterizing

Science.gov (United States)

Yuan, Yuan; Shih, F. Y.; Jing, J.; Wang, H.

2010-05-01

This paper presents a new method to extract and characterize solar filaments from H-alpha full-disk images produced by Big Bear Solar Observatory. A cascading Hough Transform method is designed to identify solar disk center location and radius. Solar disks are segmented from the background, and unbalanced illumination on the surface of solar disks is removed using polynomial surface fitting. And then a localized adaptive thresholding is employed to extract solar filament candidates. After the removal of small solar filament candidates, the remaining larger candidates are used as the seeds of region growing. The procedure of region growing not only connects broken filaments but also generate complete shape for each filament. Mathematical morphology thinning is adopted to produce the skeleton of each filament, and graph theory is used to prune branches and barbs to get the main skeleton. The length and the location of the main skeleton is characterized. The proposed method can help scientists and researches study the evolution of solar filament, for instance, to detect solar filament eruption. The presented method has already been used by Space Weather Research Lab of New Jersey Institute of Technology (http://swrl.njit.edu) to generate the solar filament online catalog using H-alpha full-disk images of Global H-alpha Network (http://swrl.njit.edu/ghn_web/).

Characterization of Campylobacter phages including analysis of host range by selected Campylobacter Penner serotypes

DEFF Research Database (Denmark)

Hansen, Vinni; Rosenquist, Hanne; Baggesen, Dorte Lau

2007-01-01

range often displayed by phages. To identify the potential of phages as a Campylobacter reducing agent we needed to determine their infectivity on a panel of isolates representing the Campylobacter strains found in broilers as well as humans. Results: In this study, Campylobacter phages were isolated...... from the intestines of broilers and ducks and from abattoir sewage. Twelve phages were investigated to determine their ability to infect the Campylobacter Penner serotypes commonly present in Danish poultry and patients with campylobacteriosis. A total of 89% of the Campylobacter jejuni strains and 14...... range of 12 Danish Campylobacter phages. Due to their ability to infect the majority of the common serotypes in Denmark we suggest the phages can become an effective agent in the effort to reduce the incidence of campylobacteriosis in Denmark. This study provides the basis for future experiments...

ORIGINAL ARTICLE: Multidrug Resistance and Phage Pattern of Staphylococcus aureus in Pyoderma Cases

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Sanjay M. Wavare

2012-01-01

Full Text Available Background: Pyoderma is common in India and other tropical countries. Staphylococcus aureus is the commonest causative agent ofpyoderma. Aims and Objectives: To know the antibiotic susceptibility and bacteriophage pattern of Staphylococcus aureus isolated from pyoderma infection. Materials and Methods: One hundred clinically diagnosed pyoderma cases were investigated bacteriologically. A total of 59 isolates of S. aureus were subjected to antibioticsusceptibility testing by Kirby Bauer’s disk diffusion method and phage typing by routine test dilution X 100 bacteriophages. Results: Most of the strains were resistant to penicillin, ampicillin and were susceptible to gentamicin, streptomycin and erythromycin. Multidrug resistance was also high among these strains. Regarding the phage types, Phage type 52 (15 strains, 96 (8 strains and 71(16strains were predominant among the typed strains (55.95% of S. aureus. The most common group was mixed phage group (17% followed by phage group I (13.55%. Conclusion: Knowledge of antibioticsusceptibility pattern is essential to give proper antibiotic therapy and avoid unnecessary medication with non-effective drugs, which may increase resistance. Gentamicin, streptomycin and erythromycin are the drugs of choice in that order. Association of phage typing and antibiotic sensitivity of S. aureus showed the predominance of phage group III with greater frequency of penicillin resistance.

Temporal symmetry of individual filaments in different spatial symmetry filaments pattern in a dielectric barrier discharge

International Nuclear Information System (INIS)

Dong, L. F.; Xiao, H.; Fan, W. L.; Yin, Z. Q.; Zhao, H. T.

2010-01-01

The temporal behavior of individual filament in different spatial symmetry filaments patterns in dielectric barrier discharge is investigated by using an optical method. A series of return maps of the discharge moments of individual filaments is given. It is found that the temporal symmetry of individual filament changes with the change of the spatial symmetry of filaments pattern as the applied voltage increases. The role of wall charges for this phenomenon is analyzed.

Magnetic helicity and active filament configuration

Science.gov (United States)

Romano, P.; Zuccarello, F.; Poedts, S.; Soenen, A.; Zuccarello, F. P.

2009-11-01

Context: The role of magnetic helicity in active filament formation and destabilization is still under debate. Aims: Although active filaments usually show a sigmoid shape and a twisted configuration before and during their eruption, it is unclear which mechanism leads to these topologies. In order to provide an observational contribution to clarify these issues, we describe a filament evolution whose characteristics seem to be directly linked to the magnetic helicity transport in corona. Methods: We applied different methods to determine the helicity sign and the chirality of the filament magnetic field. We also computed the magnetic helicity transport rate at the filament footpoints. Results: All the observational signatures provided information on the positive helicity and sinistral chirality of the flux rope containing the filament material: its forward S shape, the orientation of its barbs, the bright and dark threads at 195 Å. Moreover, the magnetic helicity transport rate at the filament footpoints showed a clear accumulation of positive helicity. Conclusions: The study of this event showed a correspondence between several signatures of the sinistral chirality of the filament and several evidences of the positive magnetic helicity of the filament magnetic field. We also found that the magnetic helicity transported along the filament footpoints showed an increase just before the change of the filament shape observed in Hα images. We argued that the photospheric regions where the filament was rooted might be the preferential ways where the magnetic helicity was injected along the filament itself and where the conditions to trigger the eruption were yielded.

Phenotypic resistance and the dynamics of bacterial escape from phage control

DEFF Research Database (Denmark)

Bull, James J.; Vegge, Christina Skovgaard; Schmerer, Matthew

2014-01-01

The canonical view of phage - bacterial interactions in dense, liquid cultures is that the phage will eliminate most of the sensitive cells; genetic resistance will then ascend to restore high bacterial densities. Yet there are various mechanisms by which bacteria may remain sensitive to phages...... mathematical models of these processes and suggest how different types of this 'phenotypic' resistance may be elucidated. We offer preliminary in vitro studies of a previously characterized E. coli model system and Campylobacter jejuni illustrating apparent phenotypic resistance. As phenotypic resistance may...

Analysis of Lactobacillus Products for Phages and Bacteriocins That Inhibit Vaginal Lactobacilli

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Lin Tao

1997-01-01

Full Text Available Objective: Bacterial vaginosis is associated with an unexplained loss of vaginal lactobacilli. Previously, we have identified certain vaginal lactobacilli-released phages that can inhibit in vitro other vaginal lactobacilli. However, there is no apparent route for phages to be transmitted among women. The purpose of this study was to identify whether certain Lactobacillus products commonly used by women release phages or bacteriocins that can inhibit vaginal lactobacilli.

Femtosecond Laser Filamentation

CERN Document Server

Chin, See Leang

2010-01-01

Femtosecond Laser Filamentation gives a comprehensive review of the physics of propagation of intense femtosecond laser pulses in optical media (principally air) and the applications and challenges of this new technique. This book presents the modern understanding of the physics of femtosecond laser pulse propagation, including unusual new effects such as the self-transformation of the pulse into a white light laser pulse, intensity clamping, the physics of multiple filamentation and competition, and how filaments’ ability to melt glass leads to wave guide writing. The potential applications of laser filamentation in atmospheric sensing and the generation of other electromagnetic pulses from the UV to the radio frequency are treated, together with possible future challenges in the excitation of super-excited states of molecules. Exciting new phenomena such as filament induced ultrafast birefringence and the excitation of molecular rotational wave packets and their multiple revivals in air (gases) will also ...

Can we determine the filament chirality by the filament footpoint location or the barb-bearing?

Science.gov (United States)

Hao, Qi; Guo, Yang; Fang, Cheng; Chen, Peng-Fei; Cao, Wen-Da

2016-01-01

We attempt to propose a method for automatically detecting the solar filament chirality and barb bearing. We first introduce the concept of an unweighted undirected graph and adopt the Dijkstra shortest path algorithm to recognize the filament spine. Then, we use the polarity inversion line (PIL) shift method for measuring the polarities on both sides of the filament, and employ the connected components labeling method to identify the barbs and calculate the angle between each barb and the spine to determine the bearing of the barbs, i.e., left or right. We test the automatic detection method with Hα filtergrams from the Big Bear Solar Observatory (BBSO) Hα archive and magnetograms observed with the Helioseismic and Magnetic Imager (HMI) on board the Solar Dynamics Observatory (SDO). Four filaments are automatically detected and illustrated to show the results. The barbs in different parts of a filament may have opposite bearings. The filaments in the southern hemisphere (northern hemisphere) mainly have left-bearing (right-bearing) barbs and positive (negative) magnetic helicity, respectively. The tested results demonstrate that our method is efficient and effective in detecting the bearing of filament barbs. It is demonstrated that the conventionally believed one-to-one correspondence between filament chirality and barb bearing is not valid. The correct detection of the filament axis chirality should be done by combining both imaging morphology and magnetic field observations.

Can we determine the filament chirality by the filament footpoint location or the barb-bearing?

International Nuclear Information System (INIS)

Hao, Qi; Guo, Yang; Fang, Cheng; Chen, Peng-Fei; Cao, Wen-Da

2016-01-01

We attempt to propose a method for automatically detecting the solar filament chirality and barb bearing. We first introduce the concept of an unweighted undirected graph and adopt the Dijkstra shortest path algorithm to recognize the filament spine. Then, we use the polarity inversion line (PIL) shift method for measuring the polarities on both sides of the filament, and employ the connected components labeling method to identify the barbs and calculate the angle between each barb and the spine to determine the bearing of the barbs, i.e., left or right. We test the automatic detection method with Hα filtergrams from the Big Bear Solar Observatory (BBSO) Hα archive and magnetograms observed with the Helioseismic and Magnetic Imager (HMI) on board the Solar Dynamics Observatory (SDO). Four filaments are automatically detected and illustrated to show the results. The barbs in different parts of a filament may have opposite bearings. The filaments in the southern hemisphere (northern hemisphere) mainly have left-bearing (right-bearing) barbs and positive (negative) magnetic helicity, respectively. The tested results demonstrate that our method is efficient and effective in detecting the bearing of filament barbs. It is demonstrated that the conventionally believed one-to-one correspondence between filament chirality and barb bearing is not valid. The correct detection of the filament axis chirality should be done by combining both imaging morphology and magnetic field observations. (paper)

Helical filaments

Energy Technology Data Exchange (ETDEWEB)

Barbieri, Nicholas; Lim, Khan; Durand, Magali; Baudelet, Matthieu; Richardson, Martin [Townes Laser Institute, CREOL—The College of Optics and Photonics, University of Central Florida, Orlando, Florida 32816 (United States); Hosseinimakarem, Zahra; Johnson, Eric [Micro-Photonics Laboratory – Center for Optical Material Science, Clemson, Anderson, South Carolina 29634 (United States)

2014-06-30

The shaping of laser-induced filamenting plasma channels into helical structures by guiding the process with a non-diffracting beam is demonstrated. This was achieved using a Bessel beam superposition to control the phase of an ultrafast laser beam possessing intensities sufficient to induce Kerr effect driven non-linear self-focusing. Several experimental methods were used to characterize the resulting beams and confirm the observed structures are laser air filaments.

Phage Types and Genotypes of Shiga Toxin-Producing Escherichia coli O157:H7 Isolates from Humans and Animals in Spain: Identification and Characterization of Two Predominating Phage Types (PT2 and PT8)

Science.gov (United States)

Mora, Azucena; Blanco, Miguel; Blanco, Jesús E.; Alonso, M. Pilar; Dhabi, Ghizlane; Thomson-Carter, Fiona; Usera, Miguel A.; Bartolomé, Rosa; Prats, Guillermo; Blanco, Jorge

2004-01-01

Phage typing and DNA macrorestriction fragment analysis by pulsed-field electrophoresis (PFGE) were used for the epidemiological subtyping of a collection of Shiga toxin-producing Escherichia coli (STEC) O157:H7 strains isolated in Spain between 1980 and 1999. Phage typing distinguished a total of 18 phage types among 171 strains isolated from different sources (67 humans, 82 bovines, 12 ovines, and 10 beef products). However, five phage types, phage type 2 (PT2; 42 strains), PT8 (33 strains), PT14 (14 strains), PT21/28 (11 strains), and PT54 (16 strains), accounted for 68% of the study isolates. PT2 and PT8 were the most frequently found among strains from both humans (51%) and bovines (46%). Interestingly, we detected a significant association between PT2 and PT14 and the presence of acute pathologies. A group of 108 of the 171 strains were analyzed by PFGE, and 53 distinct XbaI macrorestriction patterns were identified, with 38 strains exhibiting unique PFGE patterns. In contrast, phage typing identified 15 different phage types. A total of 66 phage type-PFGE subtype combinations were identified among the 108 strains. PFGE subtyping differentiated between unrelated strains that exhibited the same phage type. The most common phage type-PFGE pattern combinations were PT2-PFGE type 1 (1 human and 11 bovine strains), PT8-PFGE type 8 (2 human, 6 bovine, and 1 beef product strains), PT2-PFGE subtype 4A (1 human, 3 bovine, and 1 beef product strains). Nine (29%) of 31 human strains showed phage type-PFGE pattern combinations that were detected among the bovine strains included in this study, and 26 (38%) of 68 bovine strains produced phage type-PFGE pattern combinations observed among human strains included in this study, confirming that cattle are a major reservoir of strains pathogenic for humans. PT2 and PT8 strains formed two groups which differed from each other in their motilities, stx genotypes, PFGE patterns, and the severity of the illnesses that they caused

Identification of operator sites of the CI repressor of phage TP901-1: evolutionary link to other phages

International Nuclear Information System (INIS)

Johansen, Annette H.; Broendsted, Lone; Hammer, Karin

2003-01-01

The repressor encoded by the cI gene of the temperate Lactococcus lactis subsp. cremoris bacteriophage TP901-1 has been purified. Gel-retardation and footprinting analyses identified three palindromic operator sites (O R , O L , and O D ). The operator site O R is located between the two divergent early promoters P R and P L , O L overlaps the transcriptional start of the lytic P L promoter, and O D is located downstream of the mor gene, the first gene in the lytic gene cluster. The function of O L was verified by mutational analysis. Binding was found to be specific and cooperative. Multimeric forms of the repressor were observed, thus indicating that the repressor may bind simultaneously to all three operator sites. Inverted repeats with homology to the operator sites of TP901-1 were identified in phage genomes encoding repressors homologous to CI of TP901-1. Interestingly, the locations of these repeats on the phage genomes correspond to those found in TP901-1, indicating that the same system of cooperative repression of early phage promoters has been inherited by modular evolution

Transport of Escherichia coli phage through saturated porous media considering managed aquifer recharge.

Science.gov (United States)

Zhang, Wenjing; Li, Shuo; Wang, Shuang; Lei, Liancheng; Yu, Xipeng; Ma, Tianyi

2018-03-01

Virus is one of the most potentially harmful microorganisms in groundwater. In this paper, the effects of hydrodynamic and hydrogeochemical conditions on the transportation of the colloidal virus considering managed aquifer recharge were systematically investigated. Escherichia coli phage, vB_EcoM-ep3, has a broad host range and was able to lyse pathogenic Escherichia coli. Bacteriophage with low risk to infect human has been found extensively in the groundwater environment, so it is considered as a representative model of groundwater viruses. Laboratory studies were carried out to analyze the transport of the Escherichia coli phage under varying conditions of pH, ionic strength, cation valence, flow rate, porous media, and phosphate buffer concentration. The results indicated that decreasing the pH will increase the adsorption of Escherichia coli phage. Increasing the ionic strength, either Na + or Ca 2+ , will form negative condition for the migration of Escherichia coli phage. A comparison of different cation valence tests indicated that changes in transport and deposition were more pronounced with divalent Ca 2+ than monovalent Na + . As the flow rate increases, the release of Escherichia coli phage increases and the retention of Escherichia coli phage in the aquifer medium reduces. Changes in porous media had a significant effect on Escherichia coli phage migration. With increase of phosphate buffer concentration, the suspension stability and migration ability of Escherichia coli phage are both increased. Based on laboratory-scale column experiments, a one-dimensional transport model was established to quantitatively describe the virus transport in saturated porous medium.

Filament poisoning at typical carbon nanotube deposition conditions by hot-filament CVD

CSIR Research Space (South Africa)

Oliphant, CJ

2009-05-01

Full Text Available extensively used for the deposition of various materials, including diamond [1], polymers [2], silicon thin films [3], boron-carbon-nitride layers [4] and carbon nanotubes (CNTs) [5]. The process relies on the catalytic decomposition of precursor gases... (Ho) twice as efficient as a W filament during the deposition of microcrystalline silicon thin films [6]. Reactions between the precursor gases and the heated filament result in changes of the structural properties of the filaments; a process...

Filament wound structure and method

International Nuclear Information System (INIS)

Dritt, W.S.; Gerth, H.L.; Knight, C.E. Jr.; Pardue, R.M.

1977-01-01

A filament wound spherical structure is described comprising a plurality of filament band sets disposed about the surface of a mandrel with each band of each set formed of a continuous filament circumferentially wound about the mandrel a selected number of circuits and with each circuit of filament being wound parallel to and contiguous with an immediate previously wound circuit. Each filament band in each band set is wound at the same helix angle from the axis of revolution of the mandrel and all of the bands of each set are uniformly distributed about the mandrel circumference. The pole-to-equator wall thickness taper associated with each band set, as several contiguous band sets are wound about the mandrel starting at the poles, is accumulative as the band sets are nested to provide a complete filament wound sphere of essentially uniform thickness

Association between phage types and antimicrobial resistance among bovine Staphylococcus aureus from 10 countries

DEFF Research Database (Denmark)

Vintov, J.; Aarestrup, Frank Møller; Zinn, C. E.

2003-01-01

This study was conducted to investigate the diversity of phage types and associations between penicillin resistance and phage types among 815 Staphylococcus aureus isolates from bovine mastitis in nine European countries and USA. All isolates were examined for susceptibility to antimicrobial agents...... associated with penicillin resistance in contrast to phage group I (P = 0.0023) and phage complex-80 (P = 0.0066). This study confirms that a large number of phage types of S. aureus cause bovine mastitis, but that some types predominate. In addition, these findings could indicate that the use of penicillin...... in the bovine environment has selected for specific types of S. aureus in countries with a high frequency of resistance....

“French Phage Network”—Second Meeting Report

Science.gov (United States)

Torres-Barceló, Clara; Kaltz, Oliver; Froissart, Rémy; Gandon, Sylvain; Ginet, Nicolas; Ansaldi, Mireille

2017-01-01

The study of bacteriophages (viruses of bacteria) includes a variety of approaches, such as structural biology, genetics, ecology, and evolution, with increasingly important implications for therapeutic and industrial uses. Researchers working with phages in France have recently established a network to facilitate the exchange on complementary approaches, but also to engage new collaborations. Here, we provide a summary of the topics presented during the second meeting of the French Phage Network that took place in Marseille in November 2016. PMID:28430166

Phage “delay” towards enhancing bacterial escape from biofilms: a more comprehensive way of viewing resistance to bacteriophages

Directory of Open Access Journals (Sweden)

Stephen T. Abedon

2017-03-01

Full Text Available In exploring bacterial resistance to bacteriophages, emphasis typically is placed on those mechanisms which completely prevent phage replication. Such resistance can be detected as extensive reductions in phage ability to form plaques, that is, reduced efficiency of plating. Mechanisms include restriction-modification systems, CRISPR/Cas systems, and abortive infection systems. Alternatively, phages may be reduced in their “vigor” when infecting certain bacterial hosts, that is, with phages displaying smaller burst sizes or extended latent periods rather than being outright inactivated. It is well known, as well, that most phages poorly infect bacteria that are less metabolically active. Extracellular polymers such as biofilm matrix material also may at least slow phage penetration to bacterial surfaces. Here I suggest that such “less-robust” mechanisms of resistance to bacteriophages could serve bacteria by slowing phage propagation within bacterial biofilms, that is, delaying phage impact on multiple bacteria rather than necessarily outright preventing such impact. Related bacteria, ones that are relatively near to infected bacteria, e.g., roughly 10+ µm away, consequently may be able to escape from biofilms with greater likelihood via standard dissemination-initiating mechanisms including erosion from biofilm surfaces or seeding dispersal/central hollowing. That is, given localized areas of phage infection, so long as phage spread can be reduced in rate from initial points of contact with susceptible bacteria, then bacterial survival may be enhanced due to bacteria metaphorically “running away” to more phage-free locations. Delay mechanisms—to the extent that they are less specific in terms of what phages are targeted—collectively could represent broader bacterial strategies of phage resistance versus outright phage killing, the latter especially as require specific, evolved molecular recognition of phage presence. The

Aptamer-Phage Reporters for Ultrasensitive Lateral Flow Assays.

Science.gov (United States)

Adhikari, Meena; Strych, Ulrich; Kim, Jinsu; Goux, Heather; Dhamane, Sagar; Poongavanam, Mohan-Vivekanandan; Hagström, Anna E V; Kourentzi, Katerina; Conrad, Jacinta C; Willson, Richard C

2015-12-01

We introduce the modification of bacteriophage particles with aptamers for use as bioanalytical reporters, and demonstrate the use of these particles in ultrasensitive lateral flow assays. M13 phage displaying an in vivo biotinylatable peptide (AviTag) genetically fused to the phage tail protein pIII were used as reporter particle scaffolds, with biotinylated aptamers attached via avidin-biotin linkages, and horseradish peroxidase (HRP) reporter enzymes covalently attached to the pVIII coat protein. These modified viral nanoparticles were used in immunochromatographic sandwich assays for the direct detection of IgE and of the penicillin-binding protein from Staphylococcus aureus (PBP2a). We also developed an additional lateral flow assay for IgE, in which the analyte is sandwiched between immobilized anti-IgE antibodies and aptamer-bearing reporter phage modified with HRP. The limit of detection of this LFA was 0.13 ng/mL IgE, ∼100 times lower than those of previously reported IgE assays.

Synergistic Interaction Between Phage Therapy and Antibiotics Clears Pseudomonas Aeruginosa Infection in Endocarditis and Reduces Virulence.

Science.gov (United States)

Oechslin, Frank; Piccardi, Philippe; Mancini, Stefano; Gabard, Jérôme; Moreillon, Philippe; Entenza, José M; Resch, Gregory; Que, Yok-Ai

2017-03-01

Increasing antibiotic resistance warrants therapeutic alternatives. Here we investigated the efficacy of bacteriophage-therapy (phage) alone or combined with antibiotics against experimental endocarditis (EE) due to Pseudomonas aeruginosa, an archetype of difficult-to-treat infection. In vitro fibrin clots and rats with aortic EE were treated with an antipseudomonas phage cocktail alone or combined with ciprofloxacin. Phage pharmacology, therapeutic efficacy, and resistance were determined. In vitro, single-dose phage therapy killed 7 log colony-forming units (CFUs)/g of fibrin clots in 6 hours. Phage-resistant mutants regrew after 24 hours but were prevented by combination with ciprofloxacin (2.5 × minimum inhibitory concentration). In vivo, single-dose phage therapy killed 2.5 log CFUs/g of vegetations in 6 hours (P 6 log CFUs/g of vegetations in 6 hours and successfully treating 64% (n = 7/11) of rats. Phage-resistant mutants emerged in vitro but not in vivo, most likely because resistant mutations affected bacterial surface determinants important for infectivity (eg, the pilT and galU genes involved in pilus motility and LPS formation). Single-dose phage therapy was active against P. aeruginosa EE and highly synergistic with ciprofloxacin. Phage-resistant mutants had impaired infectivity. Phage-therapy alone or combined with antibiotics merits further clinical consideration. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America.

Combinatorial synthesis and screening of cancer cell-specific nanomedicines targeted via phage fusion proteins

Directory of Open Access Journals (Sweden)

James W. Gillespie

2015-06-01

Full Text Available Active tumor targeting of nanomedicines has recently shown significant improvements in the therapeutic activity of currently existing drug delivery systems, such as liposomal doxorubicin (Doxil/Caelyx/Lipodox. Previously, we have shown that isolated pVIII major coat proteins of the fd tet filamentous phage vector, containing cancer cell-specific peptide fusions at their N terminus, can be used as active targeting ligands in a liposomal doxorubicin delivery system in vitro and in vivo. Here, we show a novel major coat protein isolation procedure in 2-propanol that allows spontaneous incorporation of the hydrophobic protein core into preformed liposomal doxorubicin with minimal damage or drug loss while still retaining the targeting ligand exposed for cell-specific targeting. Using a panel of 12 structurally unique ligands with specificity towards breast, lung, and/or pancreatic cancer, we showed the feasibility of pVIII major coat proteins to significantly increase the throughput of targeting ligand screening in a common nanomedicine core. Phage protein-modified Lipodox samples showed an average doxorubicin recovery of 82.8% across all samples with 100% of protein incorporation in the correct orientation (N-terminus exposed. Following cytotoxicity screening in a doxorubicin-sensitive breast cancer line (MCF-7, three major groups of ligands were identified. Ligands showing the most improved cytotoxicity included: DMPGTVLP, ANGRPSMT, VNGRAEAP, and ANDVYLD showing a 25-fold improvement (p < 0.05 in toxicity. Similarly DGQYLGSQ, ETYNQPYL, and GSSEQLYL ligands with specificity towards a doxorubicin-insensitive pancreatic cancer line (PANC-1 showed significant increases in toxicity (2-fold; p < 0.05. Thus, we demonstrated proof-of-concept that pVIII major coat proteins can be screened in significantly higher throughput to identify novel ligands displaying improved therapeutic activity in a desired cancer phenotype.

The use of phage FCL-2 as an alternative to chemotherapy against columnaris disease in aquaculture

Directory of Open Access Journals (Sweden)

Elina eLaanto

2015-08-01

Full Text Available Flavobacterium columnare, the causative agent of columnaris disease in fish, causes millions of dollars of losses in the US channel catfish industry alone, not to mention aquaculture industry worldwide. Novel methods are needed for the control and treatment of bacterial diseases in aquaculture to replace traditionally used chemotherapies. A potential solution could be the use of phages, i.e., bacterial viruses, host-specific and self-enriching particles that can be can easily distributed via water flow. We examined the efficacy of phages to combat columnaris disease. A previously isolated phage, FCL-2, infecting F. columnare, was characterized by sequencing. The 47 142 bp genome of the phage had G + C content of 30.2%, and the closest similarities regarding the structural proteins were found in Cellulophaga phage phiSM. Under controlled experimental conditions, two host fish species, rainbow trout (Oncorhynchus mykiss and zebrafish (Danio rerio, were used to study the success of phage therapy to prevent F. columnare infections. The survival of both fish species was significantly higher in the presence of the phage. Hundred percent of the zebrafish and 50 % of the rainbow trout survived in the phage treatment (survival without phage 0 % and 8.3 %, respectively. Most importantly, the rainbow trout population was rescued from infection by a single addition of the phage into the water in a flow-through fish tank system. Thus, F. columnare could be used as a model system to test the benefits and risks of phage therapy on a larger scale.

Phage display as a promising approach for vaccine development

OpenAIRE

Aghebati-Maleki, Leili; Bakhshinejad, Babak; Baradaran, Behzad; Motallebnezhad, Morteza; Aghebati-Maleki, Ali; Nickho, Hamid; Yousefi, Mehdi; Majidi, Jafar

2016-01-01

Bacteriophages are specific antagonists to bacterial hosts. These viral entities have attracted growing interest as optimal vaccine delivery vehicles. Phages are well-matched for vaccine design due to being highly stable under harsh environmental conditions, simple and inexpensive large scale production, and potent adjuvant capacities. Phage vaccines have efficient immunostimulatory effects and present a high safety profile because these viruses have made a constant relationship with the mamm...

Microwave processing of ceramic oxide filaments

Energy Technology Data Exchange (ETDEWEB)

Vogt, G.J.; Katz, J.D. [Los Alamos National Laboratory, NM (United States)

1995-05-01

The objective of the microwave filament processing project is to develop microwave techniques at 2.45 GHZ to manufacture continuous ceramic oxide filaments. Microwave processing uses the volumetric absorption of microwave power in oxide filament tows to drive off process solvents, to burn out organic binders, and to sinter the dried fibers to produce flexible, high-strength ceramic filaments. The technical goal is to advance filament processing technology by microwave heating more rapidly with less energy and at a lower cost than conventional processing, but with the same quality as conventional processing. The manufacturing goal is to collaborate with the 3M Company, a US manufacturer of ceramic oxide filaments, to evaluate the technology using a prototype filament system and to transfer the microwave technology to the 3M Company.

A human gut phage catalog correlates the gut phageome with type 2 diabetes.

Science.gov (United States)

Ma, Yingfei; You, Xiaoyan; Mai, Guoqin; Tokuyasu, Taku; Liu, Chenli

2018-02-01

Substantial efforts have been made to link the gut bacterial community to many complex human diseases. Nevertheless, the gut phages are often neglected. In this study, we used multiple bioinformatic methods to catalog gut phages from whole-community metagenomic sequencing data of fecal samples collected from both type II diabetes (T2D) patients (n = 71) and normal Chinese adults (n = 74). The definition of phage operational taxonomic units (pOTUs) and identification of large phage scaffolds (n = 2567, ≥ 10 k) revealed a comprehensive human gut phageome with a substantial number of novel sequences encoding genes that were unrelated to those in known phages. Interestingly, we observed a significant increase in the number of gut phages in the T2D group and, in particular, identified 7 pOTUs specific to T2D. This finding was further validated in an independent dataset of 116 T2D and 109 control samples. Co-occurrence/exclusion analysis of the bacterial genera and pOTUs identified a complex core interaction between bacteria and phages in the human gut ecosystem, suggesting that the significant alterations of the gut phageome cannot be explained simply by co-variation with the altered bacterial hosts. Alterations in the gut bacterial community have been linked to the chronic disease T2D, but the role of gut phages therein is not well understood. This is the first study to identify a T2D-specific gut phageome, indicating the existence of other mechanisms that might govern the gut phageome in T2D patients. These findings suggest the importance of the phageome in T2D risk, which warrants further investigation.

Engineering properties of a filament-wound Kevlar 49/epoxy composite

International Nuclear Information System (INIS)

Hahn, H.T.; Chin, W.K.

1981-01-01

The effect of a flywheel service environment on transverse tension and compression, and longitudinal compression and shear properties of a filament-wound Kevlar/epoxy composite are evaluated. Shear strength and modulus were reduced by moisture desorption during preconditioning in a vacuum at 75 C, although room temperature strength and modulus increased for longitudinal compression. The desorption induced cracking of the laminate plies through increased residual stresses, which at 25 C were 15 MPa, higher than the transverse strength. The 75 C temperature caused lower strength and moduli except for longitudinal tension, and the complete test results are listed

Assessment of the microscreen phage-induction assay for screening hazardous wastes (1989)

Energy Technology Data Exchange (ETDEWEB)

Houk, V.S.; DeMarini, D.M.

1989-01-01

The Microscreen phage-induction assay, which quantitatively measures the induction of prophage Lambda in Escherichia coli WP2s(Lambda), was used to test 14 crude (unfractionated) hazardous industrial-waste samples for genotoxic activity in the presence and absence of metabolic activation. Eleven of the 14 wastes induced prophage, and induction was observed at concentrations as low as 0.4 picograms per ml. Comparisons of the mutagenic activity of these waste samples in Salmonella and their ability to induce prophage Lambda indicate that the phage-induction assay was a more-sensitive indicator of genetic damage for this group of wastes. All but one of the wastes that were mutagenic to Salmonella were detected by the phage-induction assay, and 5 wastes not mutagenic to Salmonella were genetically active in the phage assay. The enhanced ability of the phage-induction assay to detect genotoxic activity may be related to the constituents comprising these waste samples. Partial chemical characterizations of the wastes showed high concentrations of carcinogenic metals, solvents, and chlorinated compounds, most of which are detected poorly by the Salmonella assay.

Selection of staphylococcal enterotoxin B (SEB)-binding peptide using phage display technology

International Nuclear Information System (INIS)

Soykut, Esra Acar; Dudak, Fahriye Ceyda; Boyaci, Ismail Hakki

2008-01-01

In this study, peptides were selected to recognize staphylococcal enterotoxin B (SEB) which cause food intoxication and can be used as a biological war agent. By using commercial M13 phage library, single plaque isolation of 38 phages was done and binding affinities were investigated with phage-ELISA. The specificities of the selected phage clones showing high affinity to SEB were checked by using different protein molecules which can be found in food samples. Furthermore, the affinities of three selected phage clones were determined by using surface plasmon resonance (SPR) sensors. Sequence analysis was realized for three peptides showing high binding affinity to SEB and WWRPLTPESPPA, MNLHDYHRLFWY, and QHPQINQTLYRM amino acid sequences were obtained. The peptide sequence with highest affinity to SEB was synthesized with solid phase peptide synthesis technique and thermodynamic constants of the peptide-SEB interaction were determined by using isothermal titration calorimetry (ITC) and compared with those of antibody-SEB interaction. The binding constant of the peptide was determined as 4.2 ± 0.7 x 10 5 M -1 which indicates a strong binding close to that of antibody

Computational Modelling of Large Scale Phage Production Using a Two-Stage Batch Process

Directory of Open Access Journals (Sweden)

Konrad Krysiak-Baltyn

2018-04-01

Full Text Available Cost effective and scalable methods for phage production are required to meet an increasing demand for phage, as an alternative to antibiotics. Computational models can assist the optimization of such production processes. A model is developed here that can simulate the dynamics of phage population growth and production in a two-stage, self-cycling process. The model incorporates variable infection parameters as a function of bacterial growth rate and employs ordinary differential equations, allowing application to a setup with multiple reactors. The model provides simple cost estimates as a function of key operational parameters including substrate concentration, feed volume and cycling times. For the phage and bacteria pairing examined, costs and productivity varied by three orders of magnitude, with the lowest cost found to be most sensitive to the influent substrate concentration and low level setting in the first vessel. An example case study of phage production is also presented, showing how parameter values affect the production costs and estimating production times. The approach presented is flexible and can be used to optimize phage production at laboratory or factory scale by minimizing costs or maximizing productivity.

Genetic engineering of a temperate phage-based delivery system for CRISPR/Cas9 antimicrobials against Staphylococcus aureus

OpenAIRE

Park, Joo Youn; Moon, Bo Youn; Park, Juw Won; Thornton, Justin A.; Park, Yong Ho; Seo, Keun Seok

2017-01-01

Discovery of clustered, regularly interspaced, short palindromic repeats and the Cas9 RNA-guided nuclease (CRISPR/Cas9) system provides a new opportunity to create programmable gene-specific antimicrobials that are far less likely to drive resistance than conventional antibiotics. However, the practical therapeutic use of CRISPR/Cas9 is still questionable due to current shortcomings in phage-based delivery systems such as inefficient delivery, narrow host range, and potential transfer of viru...

Solar Features - Prominences and Filaments - Filaments

Data.gov (United States)

National Oceanic and Atmospheric Administration, Department of Commerce — Filaments are formed in magnetic loops that hold relatively cool, dense gas suspended above the surface of the Sun (David Hathaway/NASA)

An improved plating assay for determination of phage titer | Yang ...

African Journals Online (AJOL)

In this study, an improved plating assay was developed for detection of the number of recombinant phage Cap-T7 present in a test solution at a certain dilution point by counting the plaque forming units. The data demonstrated that the improved plating assay is fast, useful, and convenient for the determination of the phage ...

Safety analysis of a Russian phage cocktail: From MetaGenomic analysis to oral application in healthy human subjects

Energy Technology Data Exchange (ETDEWEB)

McCallin, Shawna, E-mail: semccallin@yahoo.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Alam Sarker, Shafiqul, E-mail: sasarker@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Barretto, Caroline, E-mail: Caroline.Barretto@rdls.nestle.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Sultana, Shamima, E-mail: shamima@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Berger, Bernard, E-mail: bernard.berger@rdls.nestle.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Huq, Sayeda, E-mail: sayeeda@mail.icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Krause, Lutz, E-mail: ltz.krause@gmail.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Bibiloni, Rodrigo, E-mail: Rodrigo.Bibiloni@agresearch.co.nz [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Schmitt, Bertrand, E-mail: bertrand.schmitt@rdls.nestle.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Reuteler, Gloria, E-mail: gloria.reuteler@rdls.nestle.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Brüssow, Harald, E-mail: harald.bruessow@rdls.nestle.com [Nestlé Research Centre, Nestec Ltd, Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland)

2013-09-01

Phage therapy has a long tradition in Eastern Europe, where preparations are comprised of complex phage cocktails whose compositions have not been described. We investigated the composition of a phage cocktail from the Russian pharmaceutical company Microgen targeting Escherichia coli/Proteus infections. Electron microscopy identified six phage types, with numerically T7-like phages dominating over T4-like phages. A metagenomic approach using taxonomical classification, reference mapping and de novo assembly identified 18 distinct phage types, including 7 genera of Podoviridae, 2 established and 2 proposed genera of Myoviridae, and 2 genera of Siphoviridae. De novo assembly yielded 7 contigs greater than 30 kb, including a 147-kb Myovirus genome and a 42-kb genome of a potentially new phage. Bioinformatic analysis did not reveal undesired genes and a small human volunteer trial did not associate adverse effects with oral phage exposure. - Highlights: • We analyzed the composition of a commercial Russian phage cocktail. • The cocktail consists of at least 10 different phage genera. • No undesired genes were detected. • No adverse effects were seen upon oral application in a small human clinical trial.

Safety analysis of a Russian phage cocktail: From MetaGenomic analysis to oral application in healthy human subjects

International Nuclear Information System (INIS)

McCallin, Shawna; Alam Sarker, Shafiqul; Barretto, Caroline; Sultana, Shamima; Berger, Bernard; Huq, Sayeda; Krause, Lutz; Bibiloni, Rodrigo; Schmitt, Bertrand; Reuteler, Gloria; Brüssow, Harald

2013-01-01

Phage therapy has a long tradition in Eastern Europe, where preparations are comprised of complex phage cocktails whose compositions have not been described. We investigated the composition of a phage cocktail from the Russian pharmaceutical company Microgen targeting Escherichia coli/Proteus infections. Electron microscopy identified six phage types, with numerically T7-like phages dominating over T4-like phages. A metagenomic approach using taxonomical classification, reference mapping and de novo assembly identified 18 distinct phage types, including 7 genera of Podoviridae, 2 established and 2 proposed genera of Myoviridae, and 2 genera of Siphoviridae. De novo assembly yielded 7 contigs greater than 30 kb, including a 147-kb Myovirus genome and a 42-kb genome of a potentially new phage. Bioinformatic analysis did not reveal undesired genes and a small human volunteer trial did not associate adverse effects with oral phage exposure. - Highlights: • We analyzed the composition of a commercial Russian phage cocktail. • The cocktail consists of at least 10 different phage genera. • No undesired genes were detected. • No adverse effects were seen upon oral application in a small human clinical trial

Temperate Streptococcus thermophilus phages expressing superinfection exclusion proteins of the Ltp type

Directory of Open Access Journals (Sweden)

Yahya eAli

2014-03-01

Full Text Available Lipoprotein Ltp encoded by temperate Streptococcus thermophilus phage TP-J34 is the prototype of the wide-spread family of host cell surface-exposed lipoproteins involved in superinfection exclusion. When screening for other S. thermophilus phages expressing this type of lipoprotein, three temperate phages - TP-EW, TP-DSM20617 and TP-778 - were isolated. In this communication we present the total nucleotide sequences of TP-J34 and TP-778L. For TP-EW, a phage almost identical to TP-J34, besides the ltp gene only the two regions of deviation from TP-J34 DNA were analyzed: the gene encoding the tail protein causing an assembly defect in TP-J34 and the gene encoding the lysin, which in TP-EW contains an intron. For TP-DSM20617 only the sequence of the lysogeny module containing the ltp gene was determined. The region showed high homology to the same region of TP-778. For TP-778 we could show that absence of the attR region resulted in aberrant excision of phage DNA. The amino acid sequence of mature LtpTP-EW was shown to be identical to that of mature LtpTP-J34, whereas the amino acid sequence of mature LtpTP-778 was shown to differ from mature LtpTP-J34 in eight amino acid positions. LtpTP-DSM20617 was shown to differ from LtpTP-778 in just one amino acid position. In contrast to LtpTP-J34, LtpTP-778 did not affect infection of lactococcal phage P008 instead increased activity against phage P001 was noticed.

Tungsten Filament Fire

Science.gov (United States)

Ruiz, Michael J.; Perkins, James

2016-01-01

We safely remove the outer glass bulb from an incandescent lamp and burn up the tungsten filament after the glass is removed. This demonstration dramatically illustrates the necessity of a vacuum or inert gas for the environment surrounding the tungsten filament inside the bulb. Our approach has added historical importance since the incandescent…

[Screening serum response special antibodies of U251 cell line from surface display phage antibody library].

Science.gov (United States)

Yu, Min; Tan, De-Yong; Qian, Wei; Lai, Jian-Hua; Sun, Gui-Lin

2004-05-01

U251 cell is a sensitive cell line to serum, which stops at G0 phase of cell cycle in no-serum medium, and recovers growth when the serum is added into no-serum medium. The cell can express corresponding proteins in different phase of cell cycle. Therefore it is very signification for the study of cell cycle regulation mechanism that explores these proteins. In this paper, the mouse antibody phage display library was added into the bottle in which the serum starvation U251 cells had been cultured, and the special antibody phages were absorbed. Then the absorbed antibody phages were amplified by adding E. coli TG1 and helper phage M13K07. Amplified antibody phages were added into bottle in which the serum cultured cell after serum starvation (follow named as serum recovered cells) were incubated, so that the cell absorbed the no-special antibody phages for the serum starvation cell and the special antibody phages were in supernatant. The remaining no-special antibody phages in the supernatant were discarded by repeating above program 3-4 times. The pure special antibody phages were gotten, and amplified by adding the host cell E. coli TG1 and helper phage M13K07. Then the host bacterium infected special antibody phage was spread on the plate medium with ampicillin, and the monoclonal antibody phages were gotten. Using same as above program, the monoclonal antibody phages absorbed specially for serum recovered U251 cells were obtained when the serum recovered cells instead of serum starvation cells and serum starvation cells instead of serum recovered cells. In this study, ninety-six positive monoclonal antibody phages that absorbed specially the serum starvation cells and eighty-two positive monoclonal antibody phages that absorbed specially the serum recovered cells were obtained. By using cell immunochemistry assay, two special signification antibodies were obtained. one (No.11) was the strong response in serum starvation cells, the other (No.2) was the strong

Development of a Phage Cocktail to Control Proteus mirabilis Catheter-associated Urinary Tract Infections

Science.gov (United States)

Melo, Luís D. R.; Veiga, Patrícia; Cerca, Nuno; Kropinski, Andrew M.; Almeida, Carina; Azeredo, Joana; Sillankorva, Sanna

2016-01-01

Proteus mirabilis is an enterobacterium that causes catheter-associated urinary tract infections (CAUTIs) due to its ability to colonize and form crystalline biofilms on the catheters surface. CAUTIs are very difficult to treat, since biofilm structures are highly tolerant to antibiotics. Phages have been used widely to control a diversity of bacterial species, however, a limited number of phages for P. mirabilis have been isolated and studied. Here we report the isolation of two novel virulent phages, the podovirus vB_PmiP_5460 and the myovirus vB_PmiM_5461, which are able to target, respectively, 16 of the 26 and all the Proteus strains tested in this study. Both phages have been characterized thoroughly and sequencing data revealed no traces of genes associated with lysogeny. To further evaluate the phages’ ability to prevent catheter’s colonization by Proteus, the phages adherence to silicone surfaces was assessed. Further tests in phage-coated catheters using a dynamic biofilm model simulating CAUTIs, have shown a significant reduction of P. mirabilis biofilm formation up to 168 h of catheterization. These results highlight the potential usefulness of the two isolated phages for the prevention of surface colonization by this bacterium. PMID:27446059

Survival and mutagenesis in UV-irradiated phage: Multi-hit kinetics of mutation induction and lack of indirect induction by infection with UV-irradiated phage of error-prone repair

International Nuclear Information System (INIS)

Krauss, G.; Mennigmann, H.D.; Kaplan, R.W.

1980-01-01

The paper is concerned with the question of whether Weigle-reactivation (WR) and Weigle-mutagenesis (WM) can be indirectly induced by infection with UV-irradiated phage. Experiments neither with phage lambda of Escherichia coli nor with phage kappa of Serratia marcescens show such induction. In this respect phage DNA differs from F'-DNA or Hfr-DNA; possible explanations are discussed. In both systems clear plaque mutations can also be induced by UV without irradiation of the host cells; they appear, in unirradiated and irradiated host cells, with an increase in frequency which is greater than proportional to the UV dose. It is concluded that mutation induction of phage in the unirradiated host cells is due to a low level constitutive mutagenic repair; this could either be due to 'spontaneous' induction of the mutagenic SOS function or it could be a mechanism different from this one. Host irradiation would give rise to additional activity by the induced SOS function leading to WR and WM. It is further concluded that deviation of the induction kinetics from a linear dose-dependence is not due to the necessary induction of SOS functions. (author)

Genomic Characterization of a Novel Phage Found in Black Abalone (Haliotis cracherodii) Infected with Withering Syndrome

Science.gov (United States)

Closek, C. J.; Langevin, S.; Burge, C. A.; Crosson, L.; White, S.; Friedman, C. S.

2016-02-01

Withering syndrome (WS), caused by the bacterium Candidatus Xenohaliotis californiensis, a Rickettsia-like organism (RLO), infects many species of abalone. Black abalone (Haliotis cracherodii), one of two endangered species of abalone, has experienced high population losses along the California coast due to WS. Recently, we observed reduced pathogenicity and mortality events in RLO-infected abalone when a novel bacteriophage (phage) was also present. To better understand phage-bacterium dynamics and develop more informative diagnostic tools, we sequenced the genome of the novel phage associated with the RLO responsible for WS. Metagenomic sequencing libraries were prepared with extracted genomic DNA from two experimentally infected H. cracherodii and phage sequences were enriched using hydroxyapatite chromatography normalization. Normalized libraries were individually barcoded and sequenced with Illumina MiSeq. Raw sequence reads were processed using VIrominer and de novo assembly produced one single phage-like contig (35.7Kb) from the experimentally infected abalone. This highly divergent genome had closest homology with a virus associated with abalone shriveling syndrome (SS). Of the 34 predicted ORFs, overlapping homology with the SS virus ranged from 20-72%, demonstrating the phage sequenced is genetically distinct from any known phage. The phage-like sequences represented a significant portion of the total reads sequenced ( 2 million of the 12 million paired-end reads; 17%) and we obtained 94,000X coverage across the novel phage genome. Beyond characterization of this novel phage, which appears to reduce pathogenicity of the RLO, the genome enabled us to develop quantitative PCR and in situ hybridization assays as diagnostic tools. These tools allow us to detect and quantify this phage in the endangered H. cracherodii.

Phage and bacteria support mutual diversity in a narrowing staircase of coexistence

DEFF Research Database (Denmark)

Härter, Jan Olaf Mirko; Mitarai, Namiko; Sneppen, Kim

2014-01-01

arms race will typically favor high growth rate, but a phage that infects two bacterial strains differently can occasionally eliminate the fastest growing bacteria. This context-dependent fitness allows abrupt resetting of the 'Red-Queen's race' and constrains the local diversity.......The competitive exclusion principle states that phage diversity M should not exceed bacterial diversity N. By analyzing the steady-state solutions of multistrain equations, we find a new constraint: the diversity N of bacteria living on the same resources is constrained to be M or M+1 in terms...... of the diversity of their phage predators. We quantify how the parameter space of coexistence exponentially decreases with diversity. For diversity to grow, an open or evolving ecosystem needs to climb a narrowing 'diversity staircase' by alternatingly adding new bacteria and phages. The unfolding coevolutionary...

Fine filament NbTi superconductive composite

International Nuclear Information System (INIS)

Hong, S.; Grabinsky, G.; Marancik, W.; Pattanayak, D.

1986-01-01

The large superconducting magnet for the high energy physics accelerator requires fine filament composite to minimize the field error due to the persistent current in the filaments. New concepts toward the fine filament composite and its cable fabrication are discussed. Two-stage cables of fine wire with intermediate number of filaments were introduced. The first stage was six wires cables around one and in the second stage this was used to produce a Rutherford cable. The advantage of this process is in the ease of billet fabrication since the number of filaments in a single wire is within the range of easy billet fabrication. The disadvantage is in the cable fabrication. One of the major concerns in the fabrication of fine NbTi filaments composite in a copper matrix is the intermetallic compound formation during the extrusion and heat treatment steps. The hard intermetallic particles degrade the uniformity of the filaments and reduce the critical current density. The process of using Nb barrier between the filaments and copper matrix in order to prevent this CuTi intermetallic particle formation is described

Stability of two-dimensional vorticity filaments

International Nuclear Information System (INIS)

Elhmaidi, D.; Provenzale, A.; Lili, T.; Babiano, A.

2004-01-01

We discuss the results of a numerical study on the stability of two-dimensional vorticity filaments around a circular vortex. We illustrate how the stability of the filaments depends on the balance between the strain associated with the far field of the vortex and the local vorticity of the filament, and we discuss an empirical criterion for filament stability

Impedance Measurements Could Accelerate Phage-Based Identification of Bacillus anthracis and Other Bacteria

Science.gov (United States)

2016-09-01

Impedance Measurements Could Accelerate Phage-Based Identification of Bacillus anthracis And Other Bacteria Thomas Brown, Salwa Shan, Teresa...infection can be detected as early as one hour after exposing as few as 105 CFU bacteria to the stressor. We predicted that similar responses could be used... bacteria to form confluent growth and for phage-induced plaques to appear. Techniques that permit faster detection of species-specific bacteria /phage

Unwinding motion of a twisted active region filament

Energy Technology Data Exchange (ETDEWEB)

Yan, X. L.; Xue, Z. K.; Kong, D. F. [Yunnan Observatories, Chinese Academy of Sciences, Kunming 650011 (China); Liu, J. H. [Department of Physics, Shijiazhuang University, Shijiazhuang 050035 (China); Xu, C. L. [Yunnan Normal University, Kunming 650092 (China)

2014-12-10

To better understand the structures of active region filaments and the eruption process, we study an active region filament eruption in active region NOAA 11082 in detail on 2010 June 22. Before the filament eruption, the opposite unidirectional material flows appeared in succession along the spine of the filament. The rising of the filament triggered two B-class flares at the upper part of the filament. As the bright material was injected into the filament from the sites of the flares, the filament exhibited a rapid uplift accompanying the counterclockwise rotation of the filament body. From the expansion of the filament, we can see that the filament consisted of twisted magnetic field lines. The total twist of the filament is at least 5π obtained by using a time slice method. According to the morphology change during the filament eruption, it is found that the active region filament was a twisted flux rope and its unwinding motion was like a solar tornado. We also find that there was a continuous magnetic helicity injection before and during the filament eruption. It is confirmed that magnetic helicity can be transferred from the photosphere to the filament. Using the extrapolated potential fields, the average decay index of the background magnetic fields over the filament is 0.91. Consequently, these findings imply that the mechanism of solar filament eruption could be due to the kink instability and magnetic helicity accumulation.

Filaments and clusters of galaxies

International Nuclear Information System (INIS)

Soltan, A.

1987-01-01

A statistical test to investigate filaments of galaxies is performed. Only particular form of filaments is considered, viz. filaments connecting Abell clusters of galaxies. Relative position of triplets ''cluster - field object - cluster'' is analysed. Though neither cluster sample nor field object sample are homogeneous and complete only peculiar form of selection effects could affect the present statistics. Comparison of observational data with simulations shows that less than 15 per cent of all field galaxies is concentrated in filaments connecting rich clusters. Most of the field objects used in the analysis are not normal galaxies and it is possible that this conclusion is not in conflict with apparent filaments seen in the Lick counts and in some nearby 3D maps of the galaxy distribution. 26 refs., 2 figs. (author)

The use of antibiotics to improve phage detection and enumeration by the double-layer agar technique

Directory of Open Access Journals (Sweden)

Ferreira Eugénio C

2009-07-01

Full Text Available Abstract Background The Double-Layer Agar (DLA technique is extensively used in phage research to enumerate and identify phages and to isolate mutants and new phages. Many phages form large and well-defined plaques that are easily observed so that they can be enumerated when plated by the DLA technique. However, some give rise to small and turbid plaques that are very difficult to detect and count. To overcome these problems, some authors have suggested the use of dyes to improve the contrast between the plaques and the turbid host lawns. It has been reported that some antibiotics stimulate bacteria to produce phages, resulting in an increase in final titer. Thus, antibiotics might contribute to increasing plaque size in solid media. Results Antibiotics with different mechanisms of action were tested for their ability to enhance plaque morphology without suppressing phage development. Some antibiotics increased the phage plaque surface by up to 50-fold. Conclusion This work presents a modification of the DLA technique that can be used routinely in the laboratory, leading to a more accurate enumeration of phages that would be difficult or even impossible otherwise.

Detection of sulfur mustard adducts in human callus by phage antibodies

NARCIS (Netherlands)

Bikker, F.J.; Mars-Groenendijk, R.H.; Noort, D.; Fidder, A.; Schans, G.P. van der

2007-01-01

As part of a research program to develop novel methods for diagnosis of sulfur mustard exposure in the human skin the suitability of phage display was explored. Phage display is a relative new method that enables researchers to quickly evaluate a huge range of potentially useful antibodies, thereby

Isolation and Expression of the Lysis Genes of Actinomyces naeslundii Phage Av-1

Science.gov (United States)

Delisle, Allan L.; Barcak, Gerard J.; Guo, Ming

2006-01-01

Like most gram-positive oral bacteria, Actinomyces naeslundii is resistant to salivary lysozyme and to most other lytic enzymes. We are interested in studying the lysins of phages of this important oral bacterium as potential diagnostic and therapeutic agents. To identify the Actinomyces phage genes encoding these species-specific enzymes in Escherichia coli, we constructed a new cloning vector, pAD330, that can be used to enrich for and isolate phage holin genes, which are located adjacent to the lysin genes in most phage genomes. Cloned holin insert sequences were used to design sequencing primers to identify nearby lysin genes by using whole phage DNA as the template. From partial digestions of A. naeslundii phage Av-1 genomic DNA we were able to clone, in independent experiments, inserts that complemented the defective λ holin in pAD330, as evidenced by extensive lysis after thermal induction. The DNA sequence of the inserts in these plasmids revealed that both contained the complete lysis region of Av-1, which is comprised of two holin-like genes, designated holA and holB, and an endolysin gene, designated lysA. We were able to subclone and express these genes and determine some of the functional properties of their gene products. PMID:16461656

Evolution of Filament Barbs

OpenAIRE

Liu, Rui; Xu, Yan; Wang, Haimin

2010-01-01

We present a selected few cases in which the sense of chirality of filament barbs changed within as short as hours. We investigate in detail a quiescent filament on 2003 September 10 and 11. Of its four barbs displaying such changes only one overlay a small polarity inversion line inside the EUV filament channel (EFC). No magnetic elements with magnitude above the noise level were detected at the endpoints of all barbs. In particular, a pair of barbs first approached toward and then departed ...

Viruses in the marine environment: community dynamics, phage-host interactions and genomic structure

OpenAIRE

Lara de la Casa, Elena

2014-01-01

There are an estimated 1030 viruses in the world oceans, the majority of which are phages (viruses that infect bacteria). Extensive research has demonstrated the significant influence of marine phages on microbial abundance, community structure, genetic exchange and global biogeochemical cycles. In this thesis, we contribute to increase the knowledge about the ecological role of viruses in marine systems, but also we aimed to provide a better understanding about the interactions between phage...

Phage type and sensitivity to antibiotics of Staphylococcus aureus film-forming strains isolated from airway mucosa

Directory of Open Access Journals (Sweden)

O. S. Voronkova

2014-10-01

Full Text Available Today film-forming strains of bacteria play very important role in clinical pathology. Staphylococci are ones of most dangerous of them. This bacteria can determine different pathological processes, for example, complication of airway mucosa. The ability to form a biofilm is one of the main properties of nosocomial strains. These strains should be monitored and their carriers are to be properly treated. To determine the origin of staphylococci strains we used bacteriophages from the International kit. The aim of research was to determine the phage type of staphylococci film-forming strains, that were isolated from naso-pharingial mucosa. Phage typing has been carried out for 16 film-forming strains of S. aureus. To solve this problem, we used the International phage kit by Fisher’s method. As a result, sensitivity to phages from the International kit showed 53.8% of studied strains of S. aureus. 64.3% of sensitivity strains were lysed by one of the phage, 21.4% – were by two of the phages, 14.3% – by three of the phages. Isolates were sensitive to phages: 81 – 42.9%, 75 – 35.7%, 28.6% were sensitive to phages 47 and 53. All cases of detection of sensitivity to phage 47 coincided with the ability to form biofilm. Among non-film-forming strains there was no sensitive strains for this phage. Film-forming strains resist to erythromycin (62.5%, ciprofloxacin (43.8%, gentamicin (56.3%, tetracycline (87.5%, amoxicillin (93.8%, and cefuroxime (37.5%. All cases of sensitivity to phage 47 coincided with resistance to erythromycin, amoxicillin and tetracycline. For two of these strains, we also defined resistance to gentamicin and for one of them – to ciprofloxacin. Results of research allowed to relate the bacterial cultures for determining the type. This may have implications for studying of film-forming ability, because surface structures of bacterial cell take place in this process. Belonging of an isolate to specific phage type may

Use of phages to control Vibrio splendidus infection in the juvenile sea cucumber Apostichopus japonicus.

Science.gov (United States)

Li, Zhen; Li, Xiaoyu; Zhang, Jiancheng; Wang, Xitao; Wang, Lili; Cao, Zhenhui; Xu, Yongping

2016-07-01

In the present study, we isolated 3 bacteriophages with the ability to control Vibrio splendidus, a bacterium known to cause disease in the juvenile sea cucumber. These bacteriophages were designated as vB_VspS_VS-ABTNL-1 (PVS-1), vB_VspS_VS-ABTNL-2 (PVS-2) and vB_VspS_VS-ABTNL-3 (PVS-3). The ability of the 3 phages to inhibit the growth of V. splendidus VS-ABTNL was tested in vitro using each of the 3 phages individually or in the form of a cocktail of all 3 phages in the proportion of 1:1:1. All treated cultures produced a significant (P sea cucumbers (23 ± 2 g) were randomly assigned to 1 of 6 treatments. Each treatment was housed in 3 PVC tanks (38 cm × 54 cm × 80 cm) with 20 sea cucumbers per tank. Six diets were prepared including an unsupplemented control diet, antibiotic treatment diet, 3 diets containing 1 of the 3 phages individually and a diet containing a cocktail of all 3 phages. After 60 days of feeding, all sea cucumber were challenged with V. splendidus VS-ABTNL by immersion in sea water containing a bacterial concentration of 6 × 10(6) CFU/mL for 2 days. The survival rate of sea cucumbers during the next 10 days was 18% for the unsupplemented diet, 82% for the antibiotic treatment, 82% for the phage cocktail, 65% for phage PVS-1, 58% for phage PVS-2 and 50% for phage PVS-3. There were no significant differences in weight gain, ingestion rate or feed conversion among sea cucumber fed the 4 phage treatments compared with those fed the unsupplemented diet (P > 0.05). The levels of nitric oxide synthase and acid phosphatase of sea cucumbers fed phage-containing diets were significantly (P  0.05) were detected among the 4 phage-fed treatments. An additional study was conducted in which 60 healthy sea cucumbers (23 ± 2 g) were randomly assigned to a control, an untreated group and a test group to investigate the effects of injecting phages by coelomic injection on the survival rate and enzyme activities in the coelomic fluid

Metagenomic Analysis of Therapeutic PYO Phage Cocktails from 1997 to 2014

DEFF Research Database (Denmark)

Villarroel, Julia; Larsen, Mette Voldby; Kilstrup, Mogens

2017-01-01

in the two cocktails. One of these showed no similarity to publicly available phage genomes. Representatives of phages targeting E. faecalis, E. faecium, E. coli, Proteus, P. aeruginosa and S. aureus were found in both cocktails. Finally, we estimated larger overlap of the PYO2000 cocktail to PYO97 compared...

Testing a dual-fluorescence assay to monitor the viability of filamentous cyanobacteria.

Science.gov (United States)

Johnson, Tylor J; Hildreth, Michael B; Gu, Liping; Zhou, Ruanbao; Gibbons, William R

2015-06-01

Filamentous cyanobacteria are currently being engineered to produce long-chain organic compounds, including 3rd generation biofuels. Because of their filamentous morphology, standard methods to quantify viability (e.g., plate counts) are not possible. This study investigated a dual-fluorescence assay based upon the LIVE/DEAD® BacLight™ Bacterial Viability Kit to quantify the percent viability of filamentous cyanobacteria using a microplate reader in a high throughput 96-well plate format. The manufacturer's protocol calls for an optical density normalization step to equalize the numbers of viable and non-viable cells used to generate calibration curves. Unfortunately, the isopropanol treatment used to generate non-viable cells released a blue pigment that altered absorbance readings of the non-viable cell solution, resulting in an inaccurate calibration curve. Thus we omitted this optical density normalization step, and carefully divided cell cultures into two equal fractions before the isopropanol treatment. While the resulting calibration curves had relatively high correlation coefficients, their use in various experiments resulted in viability estimates ranging from below 0% to far above 100%. We traced this to the apparent inaccuracy of the propidium iodide (PI) dye that was to stain only non-viable cells. Through further analysis via microplate reader, as well as confocal and wide-field epi-fluorescence microscopy, we observed non-specific binding of PI in viable filamentous cyanobacteria. While PI will not work for filamentous cyanobacteria, it is possible that other fluorochrome dyes could be used to selectively stain non-viable cells. This will be essential in future studies for screening mutants and optimizing photobioreactor system performance for filamentous cyanobacteria. Copyright © 2015 Elsevier B.V. All rights reserved.

Selection of phage-displayed peptides for the detection of imidacloprid in water and soil.

Science.gov (United States)

Liu, Zhiping; Liu, Jianfeng; Wang, Kai; Li, Wenhui; Shelver, Weilin L; Li, Qing X; Li, Ji; Xu, Ting

2015-09-15

Imidacloprid is the most widely used neonicotinoid insecticide in the world and shows widespread environment and human exposures. A phage clone designated L7-1 that selectively binds to imidacloprid was selected from a commercial phage display library containing linear 7-mer randomized amino acid residues. Using the clone L7-1, a competitive enzyme-linked immunosorbent assay (ELISA) for imidacloprid was developed. The half-maximum signal inhibition concentration (IC50) and the limit of detection (LOD) of the phage ELISA for imidacloprid were 96 and 2.3 ng ml(-1), respectively. This phage ELISA showed relatively low cross-reactivity with all of the tested compounds structurally similar to imidacloprid, less than 2% with the exception of 6-chloronicotinic acid, a metabolite of imidacloprid that showed 11.5%. The average recoveries of the phage ELISA for imidacloprid in water and soil samples were in the ranges of 74.6 to 86.3% and 72.5 to 93.6%, respectively. The results of the competitive phage ELISA for imidacloprid in the fortified samples agreed well with those of a high-performance liquid chromatography (HPLC) method. The simple phage-displayed peptide technology has been proven to be a convenient and efficient method for the development of an alternative format of ELISA for small molecules. Copyright © 2015 Elsevier Inc. All rights reserved.

Photoreactivation of cells and phages inactivated by UV of ecological wave-lengths

International Nuclear Information System (INIS)

Samojlova, K.A.; Yanovska, Eh.; Vizdalova, M.; Ceskoslovenska Akademie Ved, Brno. Biofysikalni Ustav)

1979-01-01

It has been found that the photoreactivity of infusoria Paramecium caudatum and bacteria Escherichia coli is high and practically similar if they are irradiated with short-wave (254 nm) and mean-wave (300-315 nm) UV radiation. The cells damaged with long-wave (315-400 nm) UV rays are not photoactivated. The latter is caused by the appearance of nonphotoreactivated damages since the phages jrradiated with the same UV rays are reactivated extremely weakly in the intact cells of bacteria (phage T7) or are not reactivated at all (phage lambdasub(c1 857))

Efficacy and Safety of a Bovine-Associated Staphylococcus aureus Phage Cocktail in a Murine Model of Mastitis

Directory of Open Access Journals (Sweden)

Koen Breyne

2017-11-01

Full Text Available Overuse of antibiotics is a major problem in the treatment of bovine mastitis, and antibiotic treatment is frequently non-curative, thus alternative treatments are necessary. The primary aim of this study was to evaluate the efficacy of a purified phage cocktail for treatment of bovine Staphylococcus aureus mastitis in a well-defined mouse model. Candidate phages were selected based on their in vitro performance and subsequently processed into an optimally composed phage cocktail. The highest scoring phages were further tested for efficacy and resistance suppression in broth and raw milk, with and without supplemental IgG. As these in vitro results displayed significant decreases in CFU, the cocktail was purified for testing in vivo. Lactating mice were intramammarily inoculated with S. aureus N305 (ATCC 29740, a clinical bovine mastitis isolate commonly used for experimental infection of dairy cows. The phage cocktail was applied via the same route 4 h post-inoculation. Treated mammary glands were graded for gross pathological appearance and excised for bacterial and phage load quantification as well as histopathology. Observation of gross macroscopic and histopathological changes and CFU quantification demonstrated that the phage cocktail treatment significantly improved mastitis pathology and decreased bacterial counts. Phage PFU quantification indicated that the tested phage cocktail treatment was able to maintain high intramammary phage titers without spreading systemically. The in vivo results complement the in vitro data and support our concept of phage therapy as an innovative alternative or supplementation therapy to antibiotics for the treatment of bovine mastitis.

Efficacy and Safety of a Bovine-Associated Staphylococcus aureus Phage Cocktail in a Murine Model of Mastitis.

Science.gov (United States)

Breyne, Koen; Honaker, Ryan W; Hobbs, Zachary; Richter, Manuela; Żaczek, Maciej; Spangler, Taylor; Steenbrugge, Jonas; Lu, Rebecca; Kinkhabwala, Anika; Marchon, Bruno; Meyer, Evelyne; Mokres, Lucia

2017-01-01

Overuse of antibiotics is a major problem in the treatment of bovine mastitis, and antibiotic treatment is frequently non-curative, thus alternative treatments are necessary. The primary aim of this study was to evaluate the efficacy of a purified phage cocktail for treatment of bovine Staphylococcus aureus mastitis in a well-defined mouse model. Candidate phages were selected based on their in vitro performance and subsequently processed into an optimally composed phage cocktail. The highest scoring phages were further tested for efficacy and resistance suppression in broth and raw milk, with and without supplemental IgG. As these in vitro results displayed significant decreases in CFU, the cocktail was purified for testing in vivo . Lactating mice were intramammarily inoculated with S. aureus N305 (ATCC 29740), a clinical bovine mastitis isolate commonly used for experimental infection of dairy cows. The phage cocktail was applied via the same route 4 h post-inoculation. Treated mammary glands were graded for gross pathological appearance and excised for bacterial and phage load quantification as well as histopathology. Observation of gross macroscopic and histopathological changes and CFU quantification demonstrated that the phage cocktail treatment significantly improved mastitis pathology and decreased bacterial counts. Phage PFU quantification indicated that the tested phage cocktail treatment was able to maintain high intramammary phage titers without spreading systemically. The in vivo results complement the in vitro data and support our concept of phage therapy as an innovative alternative or supplementation therapy to antibiotics for the treatment of bovine mastitis.

Myosin binding protein-C activates thin filaments and inhibits thick filaments in heart muscle cells.

Science.gov (United States)

Kampourakis, Thomas; Yan, Ziqian; Gautel, Mathias; Sun, Yin-Biao; Irving, Malcolm

2014-12-30

Myosin binding protein-C (MyBP-C) is a key regulatory protein in heart muscle, and mutations in the MYBPC3 gene are frequently associated with cardiomyopathy. However, the mechanism of action of MyBP-C remains poorly understood, and both activating and inhibitory effects of MyBP-C on contractility have been reported. To clarify the function of the regulatory N-terminal domains of MyBP-C, we determined their effects on the structure of thick (myosin-containing) and thin (actin-containing) filaments in intact sarcomeres of heart muscle. We used fluorescent probes on troponin C in the thin filaments and on myosin regulatory light chain in the thick filaments to monitor structural changes associated with activation of demembranated trabeculae from rat ventricle by the C1mC2 region of rat MyBP-C. C1mC2 induced larger structural changes in thin filaments than calcium activation, and these were still present when active force was blocked with blebbistatin, showing that C1mC2 directly activates the thin filaments. In contrast, structural changes in thick filaments induced by C1mC2 were smaller than those associated with calcium activation and were abolished or reversed by blebbistatin. Low concentrations of C1mC2 did not affect resting force but increased calcium sensitivity and reduced cooperativity of force and structural changes in both thin and thick filaments. These results show that the N-terminal region of MyBP-C stabilizes the ON state of thin filaments and the OFF state of thick filaments and lead to a novel hypothesis for the physiological role of MyBP-C in the regulation of cardiac contractility.

Biology of the temperate Streptococcus thermophilus bacteriophage TP-J34 and physical characterization of the phage genome

International Nuclear Information System (INIS)

Neve, Horst; Freudenberg, Wiebke; Diestel-Feddersen, Frederike; Ehlert, Regina; Heller, Knut J.

2003-01-01

The temperate Streptococcus thermophilus bacteriophage TP-J34 was identified in the lysogenic host strain J34. The majority of phage particles produced upon induction was defective and noninfectious, consisting of DNA-filled heads lacking tails. A physical map (45.6 kb) was established. Analysis of minor restriction bands of the DNA isolated from phage particles as well as the analysis of the protein pattern indicated that phage TP-J34 is a pac-type phage. This was confirmed by immunoelectron microscopy using antisera raised against virulent cos- and pac-type S. thermophilus phages. The lysogenic host J34 but not its noninducible derivate J34-12 contained phage DNA in the nonintegrated state and exhibited autolysis at elevated temperatures. Prophage-carrying strains grew homogeneously while 16 of 20 prophage-cured derivatives aggregated and sedimented rapidly. When phage TP-J34 was propagated lytically on a prophage-cured host strain, a 2.7-kb site-specific deletion occurred in the phage genome. This deletion was also identified in the prophage DNAs of relysogenized strains

Electron emitting filaments for electron discharge devices

International Nuclear Information System (INIS)

Leung, K.N.; Pincosy, P.A.; Ehlers, K.W.

1988-01-01

This patent describes an electron emitting device for use in an electron discharge system. It comprises: a filament having a pair of terminal ends, electrical supply means for supplying electrical power to the terminal ends of the filament for directly heating the filament by the passage of an electrical current along the filament between the terminal ends, the filament being substantially tapered in cross section continuously in one direction from one of its pair of terminal ends to another of its pair of terminal ends to achieve uniform heating of the filament along the length thereof by compensating for the nonuniform current along the filament due to the emission of electrons therefrom

Is phage therapy acceptable in the immunocompromised host?

Science.gov (United States)

Borysowski, Jan; Górski, Andrzej

2008-09-01

Over the last decade, bacteriophages (bacterial viruses) have emerged as the major alternative to antibiotics in the treatment of antibiotic-resistant infections. While a considerable body of evidence has accumulated for the efficacy and safety of phage therapy in immunocompetent patients, data remain relatively scarce regarding its use in the immunocompromised host. To our knowledge, the present article is the first to summarize all findings, of both experimental and clinical studies, that may be relevant to the employment of phage therapy in immunocompromised patients. The available data suggest that bacteriophages could also be an efficacious and safe therapeutic modality in such patients.

Novel phage group infecting Lactobacillus delbrueckii subsp. lactis, as revealed by genomic and proteomic analysis of bacteriophage Ldl1.

Science.gov (United States)

Casey, Eoghan; Mahony, Jennifer; Neve, Horst; Noben, Jean-Paul; Dal Bello, Fabio; van Sinderen, Douwe

2015-02-01

Ldl1 is a virulent phage infecting the dairy starter Lactobacillus delbrueckii subsp. lactis LdlS. Electron microscopy analysis revealed that this phage exhibits a large head and a long tail and bears little resemblance to other characterized phages infecting Lactobacillus delbrueckii. In vitro propagation of this phage revealed a latent period of 30 to 40 min and a burst size of 59.9 +/- 1.9 phage particles. Comparative genomic and proteomic analyses showed remarkable similarity between the genome of Ldl1 and that of Lactobacillus plantarum phage ATCC 8014-B2. The genomic and proteomic characteristics of Ldl1 demonstrate that this phage does not belong to any of the four previously recognized L. delbrueckii phage groups, necessitating the creation of a new group, called group e, thus adding to the knowledge on the diversity of phages targeting strains of this industrially important lactic acid bacterial species.

Analysis of a filament stretching rheometer

DEFF Research Database (Denmark)

Kolte, Mette Irene; Rasmussen, Henrik K.; Hassager, Ole

1996-01-01

A finite element analysis of the stretching filament rheometer of Tirtaadmadja and Sridhar (1993) is presenetd. Simulations of the stretching of a filament of the polymet test solution, fluid A, between two plates are shown.......A finite element analysis of the stretching filament rheometer of Tirtaadmadja and Sridhar (1993) is presenetd. Simulations of the stretching of a filament of the polymet test solution, fluid A, between two plates are shown....

Improved Fab presentation on phage surface with the use of molecular chaperone coplasmid system.

Science.gov (United States)

Loh, Qiuting; Leong, Siew Wen; Tye, Gee Jun; Choong, Yee Siew; Lim, Theam Soon

2015-05-15

The low presentation efficiency of Fab (fragment antigen binding) fragments during phage display is largely due to the complexity of disulphide bond formation. This can result in the presentation of Fab fragments devoid of a light chain during phage display. Here we propose the use of a coplasmid system encoding several molecular chaperones (DsbA, DsbC, FkpA, and SurA) to improve Fab packaging. A comparison was done using the Fab fragment from IgG and IgD. We found that the use of the coplasmid during phage packaging was able to improve the presentation efficiency of the Fab fragment on phage surfaces. A modified version of panning using the coplasmid system was evaluated and was successful at enriching Fab binders. Therefore, the coplasmid system would be an attractive alternative for improved Fab presentation for phage display. Copyright © 2015 Elsevier Inc. All rights reserved.

The Agricultural Antibiotic Carbadox Induces Phage-mediated Gene Transfer in Salmonella

Directory of Open Access Journals (Sweden)

Bradley L. Bearson

2014-02-01

Full Text Available Antibiotics are used for disease therapeutic or preventative effects in humans and animals, as well as for enhanced feed conversion efficiency in livestock. Antibiotics can also cause undesirable effects in microbial populations, including selection for antibiotic resistance, enhanced pathogen invasion, and stimulation of horizontal gene transfer. Carbadox is a veterinary antibiotic used in the U.S. during the starter phase of swine production for improved feed efficiency and control of swine dysentery and bacterial swine enteritis. Carbadox has been shown in vitro to induce phage-encoded Shiga toxin in Shiga toxin-producing Escherichia coli and a phage-like element transferring antibiotic resistance genes in Brachyspira hyodysenteriae, but the effect of carbadox on prophages in other bacteria is unknown. This study examined carbadox exposure on prophage induction and genetic transfer in Salmonella enterica serovar Typhimurium, a human foodborne pathogen that frequently colonizes swine without causing disease. S. Typhimurium LT2 exposed to carbadox induced prophage production, resulting in bacterial cell lysis and release of virions that were visible by electron microscopy. Carbadox induction of phage-mediated gene transfer was confirmed by monitoring the transduction of a sodCIII::neo cassette in the Fels-1 prophage from LT2 to a recipient Salmonella strain. Furthermore, carbadox frequently induced generalized transducing phages in multidrug-resistant phage type DT104 and DT120 isolates, resulting in the transfer of chromosomal and plasmid DNA that included antibiotic resistance genes. Our research indicates that exposure of Salmonella to carbadox induces prophages that can transfer virulence and antibiotic resistance genes to susceptible bacterial hosts. Carbadox-induced, phage-mediated gene transfer could serve as a contributing factor in bacterial evolution during animal production, with prophages being a reservoir for bacterial fitness

Proteomics of Filamentous Fungi

NARCIS (Netherlands)

Passel, van M.W.J.; Schaap, P.J.; Graaff, de L.H.

2013-01-01

Filamentous fungi, such as Aspergillus niger and Aspergillus oryzae traditionally have had an important role in providing enzymes and enzyme cocktails that are used in food industry. In recent years the genome sequences of many filamentous fungi have become available. This combined with

Restriction of phage T4 internal protein I mutants by a strain of Escherichia coli

International Nuclear Information System (INIS)

Black, L.W.; Abremski, K.

1974-01-01

Phage T4 internal protein I(IPI), a small (ca, 10,000 MW), basic protein injected into the host with the phage DNA, is not required for infection of most hosts, but mutants defective in IPI are restricted by at least one naturally occurring strain of Escherichia coli, CT 596 (CT). Phages lacking IPI (IPI - ) appear to inject their DNA and bind it to the membrane of CT cells as well as wild-type phage T4 does, but shutoff of host protein synthesis, initiation of T4 protein synthesis, and cell killing are abnormal in the IPI - mutant infected CT host. The injection of IPI appears to be important in allowing T4 DNA to carry out early steps involved in takeover of this host. Restriction of IPI - phage growth by CT cells appears to be due, at least in part, to a defective prophage it harbors which renders the host resistant to successful infection by phage T4 which lack IPI or rII functions. Bacteria cured of this prophage can be infected by mutants defective in these functions. The resistance of CT cells to other coliphages, and the question of T-even phage internal protein diversity are discussed. (U.S.)

The Mysterious Case of the Missing Filaments

Science.gov (United States)

Alden, C. R.

2016-12-01

Coronal Mass Ejections, or CMEs, are large solar eruptions that can have major debilitating impacts on society. Typically, these eruptions have the three following key structures: the leading edge, the empty chamber known as the cavity, and the filament which often is the brightest part of the CME. When we can see all three structures clearly with a coronagraph, it is called a classic three-part CME, also referred to as a 'lightbulb' CME. According to current knowledge, when a CME erupts, a filament should also erupt or lift off the Sun in order to have the bright center within the CME. However, we do not always see a filament erupt at the surface, and yet we still get a 'filament' within the coronagraph CME. To better understand what might be occurring with these missing filaments, we looked at three-part CMEs using the SOHO LASCO CME Catalog and filaments from the SDO AIA Filament Catalog in order to create a list of 50 CMEs without a listed filament erupting at the surface. For those CMEs without filaments in the list we closely inspected the AIA images for evidence of filament eruption. To ensure that there were no filaments past the limb of the Sun, we used data from the STEREO-A and STEREO-B spacecraft's to look at the Sun from other angles. We have found numerous events where no filament erupts from the surface, but we still see the classic three-part CME. We believe this may be due to an optical illusion occurring from the twisting of the flux rope.

The factors affecting effectiveness of treatment in phages therapy, mini review

Directory of Open Access Journals (Sweden)

Mai Huong eCHATAIN-LY

2014-02-01

Full Text Available In recent years, the use of lytic bacteriophages as antimicrobial agents controlling pathogenic bacteria has appeared as a promising new alternative strategy in the face of growing antibiotic resistance which has caused problems in many fields including medicine, veterinary medicine and aquaculture. The use of bacteriophages has numerous advantages over traditional antimicrobials. The effectiveness of phage applications in fighting against pathogenic bacteria depends on several factors such as the bacteriophages/target bacteria ratio, the mode and moment of treatment, environmental conditions (pH, temperature ..., the neutralization of phage and accessibility to target bacteria, amongst others. This report presents these factors and the challenges involved in developing phage therapy applications

Study of the phage production efficiency in the bacteria lysis processes

International Nuclear Information System (INIS)

Vidania Munoz, R. de; Garces, F.; Davila, C. A.

1979-01-01

In this work we present a search for the best production conditions of λvir andλ clear phages In E coli K12 and E coli C 6 00 infected cells respectively. By keeping fixed some parameters of the process as the bacterial and phage generation times and (he bacterial burst side, we have finder that the lysis yield is strongly dependent on the multiplicity and in a lesser degree on the infection time. It appears from the experimental results that other variables are important, as infection efficiency and approach time from phages to bacteria. We will try to describe the lysis phenomenon by a numerical model on the bases of the se experimental results. (Author) 11 refs

Selection of gonadotrophin surge attenuating factor phage antibodies by bioassay.

Science.gov (United States)

Sorsa-Leslie, Tarja; Mason, Helen D; Harris, William J; Fowler, Paul A

2005-09-26

We aimed to combine the generation of "artificial" antibodies with a rat pituitary bioassay as a new strategy to overcome 20 years of difficulties in the purification of gonadotrophin surge-attenuating factor (GnSAF). A synthetic single-chain antibody (Tomlinson J) phage display library was bio-panned with partially purified GnSAF produced by cultured human granulosa/luteal cells. The initial screening with a simple binding immunoassay resulted in 8 clones that were further screened using our in-vitro rat monolayer bioassay for GnSAF. Initially the antibodies were screened as pooled phage forms and subsequently as individual, soluble, single-chain antibody (scAbs) forms. Then, in order to improve the stability of the scAbs for immunopurification purposes, and to widen the range of labelled secondary antibodies available, these were engineered into full-length human immunoglobulins. The immunoglobulin with the highest affinity for GnSAF and a previously described rat anti-GnSAF polyclonal antiserum was then used to immunopurify bioactive GnSAF protein. The two purified preparations were electrophoresed on 1-D gels and on 7 cm 2-D gels (pH 4-7). The candidate GnSAF protein bands and spots were then excised for peptide mass mapping. Three of the scAbs recognised GnSAF bioactivity and subsequently one clone of the purified scAb-derived immunoglobulin demonstrated high affinity for GnSAF bioactivity, also binding the molecule in such as way as to block its bioactivity. When used for repeated immunopurification cycles and then Western blot, this antibody enabled the isolation of a GnSAF-bioactive protein band at around 66 kDa. Similar results were achieved using the rat anti-GnSAF polyclonal antiserum. The main candidate molecules identified from the immunopurified material by excision of 2-D gel protein spots was human serum albumin precursor and variants. This study demonstrates that the combination of bioassay and phage display technologies is a powerful tool in the

Bacteriophage T4 Infection of Stationary Phase E. coli: Life after Log from a Phage Perspective

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Elizabeth Martin Kutter

2016-09-01

Full Text Available Virtually all studies of phage infections investigate bacteria growing exponentially in rich media. In nature, however, phages largely encounter non-growing cells. Bacteria entering stationary phase often activate well-studied stress defense mechanisms that drastically alter the cell, facilitating its long-term survival. An understanding of phage-host interactions in such conditions is of major importance from both an ecological and therapeutic standpoint. Here, we show that bacteriophage T4 can efficiently bind to, infect and kill E. coli in stationary phase, both in the presence and absence of a functional stationary-phase sigma factor, and explore the response of T4-infected stationary phase cells to the addition of fresh nutrients 5 or 24 hours after that infection. An unexpected new mode of response has been identified. Hibernation mode is a persistent but reversible dormant state in which the infected cells make at least some phage enzymes, but halt phage development until appropriate nutrients become available before producing phage particles. Our evidence indicates that the block in hibernation mode occurs after the middle-mode stage of phage development; host DNA breakdown and the incorporation of the released nucleotides into phage DNA indicate that the enzymes of the nucleotide synthesizing complex, under middle-mode control, have been made and assembled into a functional state. Once fresh glucose and amino acids become available, the standard lytic infection process rapidly resumes and concentrations of up to 1011 progeny phage (an average of about 40 phage per initially-present cell are produced. All evidence is consistent with the hibernation-mode control point lying between middle mode and late mode T4 gene expression. We have also observed a scavenger response, where the infecting phage takes advantage of whatever few nutrients are available to produce small quantities of progeny within 2 to 5 hours after infection. The scavenger

Characterization of a ViI-like Phage Specific to Escherichia coli O157:H7

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Kropinski Andrew M

2011-09-01

Full Text Available Abstract Phage vB_EcoM_CBA120 (CBA120, isolated against Escherichia coli O157:H7 from a cattle feedlot, is morphologically very similar to the classic phage ViI of Salmonella enterica serovar Typhi. Until recently, little was known genetically or physiologically about the ViI-like phages, and none targeting E. coli have been described in the literature. The genome of CBA120 has been fully sequenced and is highly similar to those of both ViI and the Shigella phage AG3. The core set of structural and replication-related proteins of CBA120 are homologous to those from T-even phages, but generally are more closely related to those from T4-like phages of Vibrio, Aeromonas and cyanobacteria than those of the Enterobacteriaceae. The baseplate and method of adhesion to the host are, however, very different from those of either T4 or the cyanophages. None of the outer baseplate proteins are conserved. Instead of T4's long and short tail fibers, CBA120, like ViI, encodes tail spikes related to those normally seen on podoviruses. The 158 kb genome, like that of T4, is circularly permuted and terminally redundant, but unlike T4 CBA120 does not substitute hmdCyt for cytosine in its DNA. However, in contrast to other coliphages, CBA120 and related coliphages we have isolated cannot incorporate 3H-thymidine (3H-dThd into their DNA. Protein sequence comparisons cluster the putative "thymidylate synthase" of CBA120, ViI and AG3 much more closely with those of Delftia phage φW-14, Bacillus subtilis phage SPO1, and Pseudomonas phage YuA, all known to produce and incorporate hydroxymethyluracil (hmdUra.

Thermal Stability of RNA Phage Virus-Like Particles Displaying Foreign Peptides

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Peabody David S

2011-05-01

Full Text Available Abstract Background To be useful for genetic display of foreign peptides a viral coat protein must tolerate peptide insertions without major disruption of subunit folding and capsid assembly. The folding of the coat protein of RNA phage MS2 does not normally tolerate insertions in its AB-loop, but an engineered single-chain dimer readily accepts them as long as they are restricted to one of its two halves. Results Here we characterize the effects of peptide insertions on the thermal stabilities of MS2 virus-like particles (VLPs displaying a variety of different peptides in one AB-loop of the coat protein single-chain dimer. These particles typically denature at temperatures around 5-10°C lower than unmodified VLPs. Even so, they are generally stable up to about 50°C. VLPs of the related RNA phage PP7 are cross-linked with intersubunit disulfide bonds and are therefore significantly more stable. An AB-loop insertion also reduces the stability of PP7 VLPs, but they only begin to denature above about 70°C. Conclusions VLPs assembled from MS2 single-chain dimer coat proteins with peptide insertions in one of their AB-loops are somewhat less stable than the wild-type particle, but still resist heating up to about 50°C. Because they possess disulfide cross-links, PP7-derived VLPs provide an alternate platform with even higher stability.

Role of Intermediate Filaments in Vesicular Traffic

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Azzurra Margiotta

2016-04-01

Full Text Available Intermediate filaments are an important component of the cellular cytoskeleton. The first established role attributed to intermediate filaments was the mechanical support to cells. However, it is now clear that intermediate filaments have many different roles affecting a variety of other biological functions, such as the organization of microtubules and microfilaments, the regulation of nuclear structure and activity, the control of cell cycle and the regulation of signal transduction pathways. Furthermore, a number of intermediate filament proteins have been involved in the acquisition of tumorigenic properties. Over the last years, a strong involvement of intermediate filament proteins in the regulation of several aspects of intracellular trafficking has strongly emerged. Here, we review the functions of intermediate filaments proteins focusing mainly on the recent knowledge gained from the discovery that intermediate filaments associate with key proteins of the vesicular membrane transport machinery. In particular, we analyze the current understanding of the contribution of intermediate filaments to the endocytic pathway.

Interaction of the phage-xanthomonas campestris (Pammel) Dowson at the eletronic microscopy level, Virazole effect and radioautographic study of the phage action on the host

International Nuclear Information System (INIS)

Sittolin, I.M.

1982-04-01

A bacteriophage from the cabbage tissue infected with Xanthomonas campestris is described. The infection process is studied through a negative staining technique (PTA) and ultrathin section. The effect of Virazole, an antivirus agent, is tested. Radioautography showed that the phage presented a reasonable domain on the bacterial host genome since the beginning of the treatment. Sorological reactions indicated the induction of specific antibodies for the phage. (M.A.C.) [pt

The genome and structural proteome of YuA, a new Pseudomonas aeruginosa phage resembling M6.

Science.gov (United States)

Ceyssens, Pieter-Jan; Mesyanzhinov, Vadim; Sykilinda, Nina; Briers, Yves; Roucourt, Bart; Lavigne, Rob; Robben, Johan; Domashin, Artem; Miroshnikov, Konstantin; Volckaert, Guido; Hertveldt, Kirsten

2008-02-01

Pseudomonas aeruginosa phage YuA (Siphoviridae) was isolated from a pond near Moscow, Russia. It has an elongated head, encapsulating a circularly permuted genome of 58,663 bp, and a flexible, noncontractile tail, which is terminally and subterminally decorated with short fibers. The YuA genome is neither Mu- nor lambda-like and encodes 78 gene products that cluster in three major regions involved in (i) DNA metabolism and replication, (ii) host interaction, and (iii) phage particle formation and host lysis. At the protein level, YuA displays significant homology with phages M6, phiJL001, 73, B3, DMS3, and D3112. Eighteen YuA proteins were identified as part of the phage particle by mass spectrometry analysis. Five different bacterial promoters were experimentally identified using a promoter trap assay, three of which have a sigma54-specific binding site and regulate transcription in the genome region involved in phage particle formation and host lysis. The dependency of these promoters on the host sigma54 factor was confirmed by analysis of an rpoN mutant strain of P. aeruginosa PAO1. At the DNA level, YuA is 91% identical to the recently (July 2007) annotated phage M6 of the Lindberg typing set. Despite this level of DNA homology throughout the genome, both phages combined have 15 unique genes that do not occur in the other phage. The genome organization of both phages differs substantially from those of the other known Pseudomonas-infecting Siphoviridae, delineating them as a distinct genus within this family.

Soliton on thin vortex filament

International Nuclear Information System (INIS)

Konno, Kimiaki; Mituhashi, Masahiko; Ichikawa, Y.H.

1990-12-01

Showing that one of the equations found by Wadati, Konno and Ichikawa is equivalent to the equation of motion of a thin vortex filament, we investigate solitons on the vortex filament. N vortex soliton solution is given in terms of the inverse scattering method. We examine two soliton collision processes on the filament. Our analysis provides the theoretical foundation of two soliton collision processes observed numerically by Aref and Flinchem. (author)

X-ray inactivation and reactivation characteristics of the phage 'kappa'

International Nuclear Information System (INIS)

Bhattacharyya, S.C.; Samad, S.A.; Mandal, J.C.; Chatterjee, S.N.

1991-01-01

Vibrio cholerae temperate phage 'kappa' was inactivated by X-ray (60 kV) in a dose dependent manner, the inactivation dose leading to 37% survival (D 37 ) in PBS, pH 7.4 being 0.36 kGy. The phages were significantly protected against X-ray irradiation when histidine or cysteine or both were present in PBS or when phages were irradiated in nutrient broth. The maximum protection was offered when histidine (10.0 nM) and cysteine (10.0 nM) were both present in PBS (dose enhancement factor being 4.17). The X-irradiated 'kappa' phages also underwent a small but significant Weigle reactivation and also Weigle mutagenesis in the UV-irradiated V. cholerae host H218Sm r . The Weigle factor (WF) or the frequency of clear plaque mutants increased with increasing UV dose, attained a maximum at the UV dose of 2.4 Jm -2 and thereafter decreased gradually with further increase of UV dose. The X-ray dose (D)-survival (S) curves could be empirically described by the equation S=exp-(aD+bD 2 ) where 'a' and 'b' are constants depending on the irradiation conditions and good agreement between the theoretical curves and experimental data was obtained. (author). 1 5 refs., 2 fig., 1 tab

The extracellular phage-host interactions involved in the bacteriophage LL-H infection of Lactobacillus delbrueckii ssp. lactis ATCC 15808.

Science.gov (United States)

Munsch-Alatossava, Patricia; Alatossava, Tapani

2013-12-24

The complete genome sequence of Lactobacillus bacteriophage LL-H was determined in 1996. Accordingly, LL-H has been used as a model phage for the infection of dairy Lactobacillus, specifically for thermophilic Lactobacillus delbrueckii ssp. lactis host strains, such as ATCC 15808. One of the major goals of phage LL-H research consisted of the characterization of the first phage-host interactions at the level of phage adsorption and phage DNA injection steps to determine effective and practical methods to minimize the risks associated with the appearance and attack of phages in the manufacture of yogurt, and Swiss or Italian hard type cheeses, which typically use thermophilic lactic acid bacteria starter cultures containing L. delbrueckii strains among others. This mini review article summarizes the present data concerning (i) the special features, particle structure, and components of phage LL-H and (ii) the structure and properties of lipoteichoic acids (LTAs), which are the phage LL-H receptor components of L. delbrueckii ssp. lactis host strains. Moreover, a model of the first, extracellular, phage-host interactions for the infection of L. delbrueckii ssp. lactis ATCC 15808 by phage LL-H is presented and further discussed.

The extracellular phage-host interactions involved in the bacteriophage LL-H infection of Lactobacillus delbrueckii ssp. lactis ATCC 15808

Directory of Open Access Journals (Sweden)

Patricia eMunsch-Alatossava

2013-12-01

Full Text Available The complete genome sequence of Lactobacillus bacteriophage LL-H was determined in 1996. Accordingly, LL-H has been used as a model phage for the infection of dairy Lactobacillus, specifically for thermophilic Lb. delbrueckii ssp. lactis host strains, such as ATCC 15808. One of the major goals of phage LL-H research consisted of the characterization of the the first phage-host interactions at the level of phage adsorption and phage DNA injection steps to determine effective and practical methods to minimise the risks associated with the appearance and attack of phages in the manufacture of yoghurt, and Swiss or Italian type hard cheeses, which typically use thermophilic LAB starter cultures containing Lb. delbrueckii strains among others. This mini review article summarises the present data concerning (i the special features, particle structure and components of phage LL-H and (ii the structure and properties of lipoteichoic acids (LTAs, which are the phage LL-H receptor components of Lb. delbrueckii ssp. lactis host strains. Moreover, a model of the first, extracellular, phage-host interactions for the infection of Lb. delbrueckii ssp. lactis ATCC 15808 by phage LL-H is presented and further discussed.

Striation and convection in penumbral filaments

NARCIS (Netherlands)

Spruit, H.C.; Scharmer, G.B.; Löfdahl, M.G.

2010-01-01

Observations with the 1-m Swedish Solar Telescope of the flows seen in penumbral filaments are presented. Time sequences of bright filaments show overturning motions strikingly similar to those seen along the walls of small isolated structures in the active regions. The filaments show outward

Boolean gates on actin filaments

International Nuclear Information System (INIS)

Siccardi, Stefano; Tuszynski, Jack A.; Adamatzky, Andrew

2016-01-01

Actin is a globular protein which forms long polar filaments in the eukaryotic cytoskeleton. Actin networks play a key role in cell mechanics and cell motility. They have also been implicated in information transmission and processing, memory and learning in neuronal cells. The actin filaments have been shown to support propagation of voltage pulses. Here we apply a coupled nonlinear transmission line model of actin filaments to study interactions between voltage pulses. To represent digital information we assign a logical TRUTH value to the presence of a voltage pulse in a given location of the actin filament, and FALSE to the pulse's absence, so that information flows along the filament with pulse transmission. When two pulses, representing Boolean values of input variables, interact, then they can facilitate or inhibit further propagation of each other. We explore this phenomenon to construct Boolean logical gates and a one-bit half-adder with interacting voltage pulses. We discuss implications of these findings on cellular process and technological applications. - Highlights: • We simulate interaction between voltage pulses using on actin filaments. • We use a coupled nonlinear transmission line model. • We design Boolean logical gates via interactions between the voltage pulses. • We construct one-bit half-adder with interacting voltage pulses.

Boolean gates on actin filaments

Energy Technology Data Exchange (ETDEWEB)

Siccardi, Stefano, E-mail: ssiccardi@2ssas.it [The Unconventional Computing Centre, University of the West of England, Bristol (United Kingdom); Tuszynski, Jack A., E-mail: jackt@ualberta.ca [Department of Oncology, University of Alberta, Edmonton, Alberta (Canada); Adamatzky, Andrew, E-mail: andrew.adamatzky@uwe.ac.uk [The Unconventional Computing Centre, University of the West of England, Bristol (United Kingdom)

2016-01-08

Actin is a globular protein which forms long polar filaments in the eukaryotic cytoskeleton. Actin networks play a key role in cell mechanics and cell motility. They have also been implicated in information transmission and processing, memory and learning in neuronal cells. The actin filaments have been shown to support propagation of voltage pulses. Here we apply a coupled nonlinear transmission line model of actin filaments to study interactions between voltage pulses. To represent digital information we assign a logical TRUTH value to the presence of a voltage pulse in a given location of the actin filament, and FALSE to the pulse's absence, so that information flows along the filament with pulse transmission. When two pulses, representing Boolean values of input variables, interact, then they can facilitate or inhibit further propagation of each other. We explore this phenomenon to construct Boolean logical gates and a one-bit half-adder with interacting voltage pulses. We discuss implications of these findings on cellular process and technological applications. - Highlights: • We simulate interaction between voltage pulses using on actin filaments. • We use a coupled nonlinear transmission line model. • We design Boolean logical gates via interactions between the voltage pulses. • We construct one-bit half-adder with interacting voltage pulses.

Phage Lambda P Protein: Trans-Activation, Inhibition Phenotypes and their Suppression

Science.gov (United States)

Hayes, Sidney; Erker, Craig; Horbay, Monique A.; Marciniuk, Kristen; Wang, Wen; Hayes, Connie

2013-01-01

The initiation of bacteriophage λ replication depends upon interactions between the oriλ DNA site, phage proteins O and P, and E. coli host replication proteins. P exhibits a high affinity for DnaB, the major replicative helicase for unwinding double stranded DNA. The concept of P-lethality relates to the hypothesis that P can sequester DnaB and in turn prevent cellular replication initiation from oriC. Alternatively, it was suggested that P-lethality does not involve an interaction between P and DnaB, but is targeted to DnaA. P-lethality is assessed by examining host cells for transformation by ColE1-type plasmids that can express P, and the absence of transformants is attributed to a lethal effect of P expression. The plasmid we employed enabled conditional expression of P, where under permissive conditions, cells were efficiently transformed. We observed that ColE1 replication and plasmid establishment upon transformation is extremely sensitive to P, and distinguish this effect from P-lethality directed to cells. We show that alleles of dnaB protect the variant cells from P expression. P-dependent cellular filamentation arose in ΔrecA or lexA[Ind-] cells, defective for SOS induction. Replication propagation and restart could represent additional targets for P interference of E. coli replication, beyond the oriC-dependent initiation step. PMID:23389467

Phage-resistance linked to cell heterogeneity in the commercial strain Lactobacillus delbrueckii subsp. lactis Ab1.

Science.gov (United States)

Suárez, Viviana B; Maciel, Natalia; Guglielmotti, Daniela; Zago, Miriam; Giraffa, Giorgio; Reinheimer, Jorge

2008-12-10

The aim of this work was to study the relationship between the cell morphological heterogeneity and the phage-resistance in the commercial strain Lactobacillus delbrueckii subsp. lactis Ab1. Two morphological variants (named C and T) were isolated from this strain. Phage-resistant derivatives were isolated from them and the percentage of occurrence of confirmed phage-resistant cells was 0.001% of the total cellular population. Within these phage-resistant cell derivatives there were T (3 out of 4 total isolates) and C (1 out of 4 total isolates) variants. The study of some technological properties (e.g. proteolytic and acidifying activities) demonstrated that most of phage-resistant derivatives were not as good as the parental strain. However, for one derivative (a T variant), the technological properties were better than those of the parental strain. On the other hand, it was possible to determinate that the system of phage-resistance in the T variants was interference in adsorption step, with adsorption rates M.

Thermal-Stability and Reconstitution Ability of Listeria Phages P100 and A511

Directory of Open Access Journals (Sweden)

Hanie Ahmadi

2017-12-01

Full Text Available The study evaluated the thermal-stability of Listeria phages P100 and A511 at temperatures simulating the preparation of ready-to-eat meats. The phage infectivity after heating to 71°C and holding for a minimum of 30 s, before eventually cooling to 4°C were examined. Higher temperatures of 75, 80, and 85°C were also tested to evaluate their effect on phages thermal-stability. This study found that despite minor differences in the amino acid sequences of their structural proteins, the two phages responded differently to high temperatures. P100 activity declined at least 10 log (PFU mL-1 with exposure to 71°C (30 s and falling below the limit of detection (1 log PFU mL-1 while, A511 dropped from 108 to 105 PFU mL-1. Cooling resulted in partial reconstitution of P100 phage particles to 103 PFU mL-1. Exposure to 75°C (30 s abolished A511 activity (8 log PFU mL-1 and both phages showed reconstitution during cooling phase after exposure to 75°C. P100 exhibited reconstitution after treatment at 80°C (30 s, conversely A511 showed no reconstitution activity. Heating P100 to 85°C abolished the reconstitution potential. Substantial differences were found in thermal-stability and reconstitution of the examined phages showing A511 to be more thermo-stable than P100, while P100 exhibited reconstitution during cooling after treatment at 80°C which was absent in A511. The differences in predicted melting temperatures of structural proteins of P100 and A511 were consistent with the observed differences in thermal stability and morphological changes observed with transmission electron microscopy.

Solar filament material oscillations and drainage before eruption

International Nuclear Information System (INIS)

Bi, Yi; Jiang, Yunchun; Yang, Jiayan; Hong, Junchao; Li, Haidong; Yang, Dan; Yang, Bo

2014-01-01

Both large-amplitude longitudinal (LAL) oscillations and material drainage in a solar filament are associated with the flow of material along the filament axis, often followed by an eruption. However, the relationship between these two motions and a subsequent eruption event is poorly understood. We analyze a filament eruption using EUV imaging data captured by the Atmospheric Imaging Array on board the Solar Dynamics Observatory and the Hα images from the Global Oscillation Network Group. Hours before the eruption, the filament was activated, with one of its legs undergoing a slow rising motion. The asymmetric activation inclined the filament relative to the solar surface. After the active phase, LAL oscillations were observed in the inclined filament. The oscillation period increased slightly over time, which may suggest that the magnetic fields supporting the filament evolve to be flatter during the slow rising phase. After the oscillations, a significant amount of filament material was drained toward one filament endpoint, followed immediately by the violent eruption of the filament. The material drainage may further support the change in magnetic topology prior to the eruption. Moreover, we suggest that the filament material drainage could play a role in the transition from a slow to a fast rise of the erupting filament.

Prediction of Solar Eruptions Using Filament Metadata

Science.gov (United States)

Aggarwal, Ashna; Schanche, Nicole; Reeves, Katharine K.; Kempton, Dustin; Angryk, Rafal

2018-05-01

We perform a statistical analysis of erupting and non-erupting solar filaments to determine the properties related to the eruption potential. In order to perform this study, we correlate filament eruptions documented in the Heliophysics Event Knowledgebase (HEK) with HEK filaments that have been grouped together using a spatiotemporal tracking algorithm. The HEK provides metadata about each filament instance, including values for length, area, tilt, and chirality. We add additional metadata properties such as the distance from the nearest active region and the magnetic field decay index. We compare trends in the metadata from erupting and non-erupting filament tracks to discover which properties present signs of an eruption. We find that a change in filament length over time is the most important factor in discriminating between erupting and non-erupting filament tracks, with erupting tracks being more likely to have decreasing length. We attempt to find an ensemble of predictive filament metadata using a Random Forest Classifier approach, but find the probability of correctly predicting an eruption with the current metadata is only slightly better than chance.

Recent Trends in Salmonella Outbreaks and Emerging Technology for Biocontrol of Salmonella Using Phages in Foods: A Review.

Science.gov (United States)

Oh, Jun-Hyun; Park, Mi-Kyung

2017-12-28

Salmonella is one of the principal causes of foodborne outbreaks. As traditional control methods have shown less efficacy against emerging Salmonella serotypes or antimicrobialresistant Salmonella , new approaches have been attempted. The use of lytic phages for the biocontrol of Salmonella in the food industry has become an attractive method owing to the many advantages offered by the use of phages as biocontrol agents. Phages are natural alternatives to traditional antimicrobial agents; they have proven effective in the control of bacterial pathogens in the food industry, which has led to the development of different phage products. The treatment with specific phages in the food industry can prevent the decay of products and the spread of bacterial diseases, and ultimately promotes safe environments for animal and plant food production, processing, and handling. After an extensive investigation of the current literature, this review focuses predominantly on the efficacy of phages for the successful control of Salmonella spp. in foods. This review also addresses the current knowledge on the pathogenic characteristics of Salmonella , the prevalence of emerging Salmonella outbreaks, the isolation and characterization of Salmonella -specific phages, the effectiveness of Salmonella -specific phages as biocontrol agents, and the prospective use of Salmonella -specific phages in the food industry.

Association between phage types and antimicrobial resistance among bovine isolates of Staphylococcus aureus in 10 countries

DEFF Research Database (Denmark)

Vintov, J.; Aarestrup, Frank Møller; Zinn, C. E.

2003-01-01

This study was conducted to investigate the diversity of phage types and associations between penicillin resistance and phage types among 815 Staphylococcus aureus isolates from bovine mastitis in nine European countries and USA. All isolates were examined for susceptibility to antimicrobial agents...... associated with penicillin resistance in contrast to phage group I (P = 0.0023) and phage complex-80 (P = 0.0066). This study confirms that a large number of phage types of S. aureus cause bovine mastitis, but that some types predominate. In addition, these findings could indicate that the use of penicillin...... in the bovine environment has selected for specific types of S. aureus in countries with a high frequency of resistance. (C) 2003 Elsevier B.V. All rights reserved....

A study of Salmonella typhi isolated in Suez Canal area. Biotyping, phage typing and colicinogenic property.

Science.gov (United States)

Shoeb, S; Khalifa, I; el Daly, O; Heiba, A; Farmer, J; Brenner, F; el Batawi, Y

1989-01-01

In this work a total of 82 strains of Salmonella typhi were isolated from Egyptian patients diagnosed as quiry enteric fever. These cases were from Ismalia, Suez and port Said Areas. The strains fell in 16 phage types. Phage types N, 40, E1, and degraded Vi were the commonest phage type in Ismailia, while phage types degraded Vi and C1 were the commonest in Port Said. Phage types Di-N, degraded Vi, A and C1 were the commonest in Suez. Chemotyping of Salmonella typhi showed that the majority of the strains belonged to chemotype I (82%), and the rest belonged to chemotype II (18%). Colicin production was negative and all the strains were susceptible to the currently used antibiotics.

Two Novel Myoviruses from the North of Iraq Reveal Insights into Clostridium difficile Phage Diversity and Biology

Directory of Open Access Journals (Sweden)

Srwa J. Rashid

2016-11-01

Full Text Available Bacteriophages (phages are increasingly being explored as therapeutic agents to combat bacterial diseases, including Clostridium difficile infections. Therapeutic phages need to be able to efficiently target and kill a wide range of clinically relevant strains. While many phage groups have yet to be investigated in detail, those with new and useful properties can potentially be identified when phages from newly studied geographies are characterised. Here, we report the isolation of C. difficile phages from soil samples from the north of Iraq. Two myoviruses, CDKM15 and CDKM9, were selected for detailed sequence analysis on the basis of their broad and potentially useful host range. CDKM9 infects 25/80 strains from 12/20 C. difficile ribotypes, and CDKM15 infects 20/80 strains from 9/20 ribotypes. Both phages can infect the clinically relevant ribotypes R027 and R001. Phylogenetic analysis based on whole genome sequencing revealed that the phages are genetically distinct from each other but closely related to other long-tailed myoviruses. A comparative genomic analysis revealed key differences in the genes predicted to encode for proteins involved in bacterial infection. Notably, CDKM15 carries a clustered regularly interspaced short palindromic repeat (CRISPR array with spacers that are homologous to sequences in the CDKM9 genome and of phages from diverse localities. The findings presented suggest a possible shared evolutionary past for these phages and provides evidence of their widespread dispersal.

Temperature distributions of a conductively heated filament

International Nuclear Information System (INIS)

Tamura, Koji; Ohba, Hironori; Shibata, Takemasa

1999-07-01

Temperature distributions of a heated filament were measured. A W-Re(5%) filament (0.25 mm in diameter, 24.7 mm in length) was conductively heated by currents between 5A and 7A with a DC power supply, and the surface of the filament was imaged with a charge coupled device (CCD) camera through a monochromatic filter. The spectral radiation intensity at the filament center region was almost uniform. Since the temperature distribution was also uniform and the energy loss by thermal conduction was negligible, temperature in this region was determined from the energy balance between applied power and radiation loss. Temperature distribution of the filament was determined based on the Planck's law of radiation from the spectral radiation intensity ratio of the filament surface using obtained temperature as a reference. It was found that temperature distribution of a filament was easily measured by this method. (author)

Comparative genomics and functional analysis of the 936 group of lactococcal Siphoviridae phages

NARCIS (Netherlands)

Murphy, James; Bottacini, Francesca; Mahony, Jennifer; Kelleher, Philip; Neve, Horst; Zomer, Aldert; Nauta, Arjen; van Sinderen, Douwe

2016-01-01

Genome sequencing and comparative analysis of bacteriophage collections has greatly enhanced our understanding regarding their prevalence, phage-host interactions as well as the overall biodiversity of their genomes. This knowledge is very relevant to phages infecting Lactococcus lactis, since they

Measuring Filament Orientation: A New Quantitative, Local Approach

Energy Technology Data Exchange (ETDEWEB)

Green, C.-E.; Cunningham, M. R.; Jones, P. A. [School of Physics, University of New South Wales, Sydney, NSW, 2052 (Australia); Dawson, J. R. [CSIRO Astronomy and Space Science, Australia Telescope National Facility, P.O. Box 76, Epping, NSW 1710 (Australia); Novak, G. [Center for Interdisciplinary Exploration and Research in Astrophysics (CIERA) and Department of Physics and Astronomy, Northwestern University, 2145 Sheridan Road, Evanston, IL 60208 (United States); Fissel, L. M. [National Radio Astronomy Observatory (NRAO), 520 Edgemont Road, Charlottesville, VA, 22903 (United States)

2017-09-01

The relative orientation between filamentary structures in molecular clouds and the ambient magnetic field provides insight into filament formation and stability. To calculate the relative orientation, a measurement of filament orientation is first required. We propose a new method to calculate the orientation of the one-pixel-wide filament skeleton that is output by filament identification algorithms such as filfinder. We derive the local filament orientation from the direction of the intensity gradient in the skeleton image using the Sobel filter and a few simple post-processing steps. We call this the “Sobel-gradient method.” The resulting filament orientation map can be compared quantitatively on a local scale with the magnetic field orientation map to then find the relative orientation of the filament with respect to the magnetic field at each point along the filament. It can also be used for constructing radial profiles for filament width fitting. The proposed method facilitates automation in analyses of filament skeletons, which is imperative in this era of “big data.”.

Measuring Filament Orientation: A New Quantitative, Local Approach

Science.gov (United States)

Green, C.-E.; Dawson, J. R.; Cunningham, M. R.; Jones, P. A.; Novak, G.; Fissel, L. M.

2017-09-01

The relative orientation between filamentary structures in molecular clouds and the ambient magnetic field provides insight into filament formation and stability. To calculate the relative orientation, a measurement of filament orientation is first required. We propose a new method to calculate the orientation of the one-pixel-wide filament skeleton that is output by filament identification algorithms such as filfinder. We derive the local filament orientation from the direction of the intensity gradient in the skeleton image using the Sobel filter and a few simple post-processing steps. We call this the “Sobel-gradient method.” The resulting filament orientation map can be compared quantitatively on a local scale with the magnetic field orientation map to then find the relative orientation of the filament with respect to the magnetic field at each point along the filament. It can also be used for constructing radial profiles for filament width fitting. The proposed method facilitates automation in analyses of filament skeletons, which is imperative in this era of “big data.”

Measuring Filament Orientation: A New Quantitative, Local Approach

International Nuclear Information System (INIS)

Green, C.-E.; Cunningham, M. R.; Jones, P. A.; Dawson, J. R.; Novak, G.; Fissel, L. M.

2017-01-01

The relative orientation between filamentary structures in molecular clouds and the ambient magnetic field provides insight into filament formation and stability. To calculate the relative orientation, a measurement of filament orientation is first required. We propose a new method to calculate the orientation of the one-pixel-wide filament skeleton that is output by filament identification algorithms such as filfinder. We derive the local filament orientation from the direction of the intensity gradient in the skeleton image using the Sobel filter and a few simple post-processing steps. We call this the “Sobel-gradient method.” The resulting filament orientation map can be compared quantitatively on a local scale with the magnetic field orientation map to then find the relative orientation of the filament with respect to the magnetic field at each point along the filament. It can also be used for constructing radial profiles for filament width fitting. The proposed method facilitates automation in analyses of filament skeletons, which is imperative in this era of “big data.”

An efficient method for isolating antibody fragments against small peptides by antibody phage display

DEFF Research Database (Denmark)

Duan, Zhi; Siegumfeldt, Henrik

2010-01-01

We generated monoclonal scFv (single chain variable fragment) antibodies from an antibody phage display library towards three small synthetic peptides derived from the sequence of s1-casein. Key difficulties for selection of scFv-phages against small peptides were addressed. Small peptides do....... The scFvs were sequenced and characterized, and specificity was characterized by ELISA. The methods developed in this study are universally applicable for antibody phage display to efficiently produce antibody fragments against small peptides....

Solar Features - Prominences and Filaments

Data.gov (United States)

National Oceanic and Atmospheric Administration, Department of Commerce — Prominences and filaments are two manifestations of the same phenomenon. Both prominences and filaments are features formed above the chromosphere by cool dense...

Dynamics and mechanics of motor-filament systems

Science.gov (United States)

Kruse, K.; Jülicher, F.

2006-08-01

Motivated by the cytoskeleton of eukaryotic cells, we develop a general framework for describing the large-scale dynamics of an active filament network. In the cytoskeleton, active cross-links are formed by motor proteins that are able to induce relative motion between filaments. Starting from pair-wise interactions of filaments via such active processes, our framework is based on momentum conservation and an analysis of the momentum flux. This allows us to calculate the stresses in the filament network generated by the action of motor proteins. We derive effective theories for the filament dynamics which can be related to continuum theories of active polar gels. As an example, we discuss the stability of homogenous isotropic filament distributions in two spatial dimensions.

[Construction of human phage antibody library and screening for human monoclonal antibodies of amylin].

Science.gov (United States)

Gong, Qian; Li, Chang-ying; Chang, Ji-wu; Zhu, Tie-hong

2012-06-01

To screen monoclonal antibodies to amylin from a constructed human phage antibody library and identify their antigenic specificity and combining activities. The heavy chain Fd fragment and light chain of human immunoglobulin genes were amplified from peripheral blood lymphocytes of healthy donors using RT-PCR, and then inserted into phagemid pComb3XSS to generate a human phage antibody library. The insertion of light chain or heavy chain Fd genes were identified by PCR after the digestion of Sac I, Xba I, Xho Iand Spe I. One of positive clones was analyzed by DNA sequencing. The specific anti-amylin clones were screened from antibody library against human amylin antigens and then the positive clones were determined by Phage-ELISA analysis. A Fab phage antibody library with 0.8×10(8); members was constructed with the efficacy of about 70%. DNA sequence analysis indicated V(H); gene belonged to V(H);3 gene family and V(λ); gene belonged to the V(λ); gene family. Using human amylin as panning antigen, specific anti-amylin Fab antibodies were enriched by screening the library for three times. Phage-ELISA assay showed the positive clones had very good specificity to amylin antigen. The successful construction of a phage antibody library and the identification of anti-amylin Fab antibodies provide a basis for further study and preparation of human anti-amylin antibodies.

TRANSDUCTION OF BACILLUS LICHENIFORMIS AND BACILLUS SUBTILIS BY EACH OF TWO PHAGES1

Science.gov (United States)

Taylor, Martha J.; Thorne, Curtis B.

1963-01-01

Taylor, Martha J. (U.S. Army Biological Laboratories, Fort Detrick, Frederick, Md.) and Curtis B. Thorne. Transduction of Bacillus licheniformis and Bacillus subtilis by each of two phages. J. Bacteriol. 86:452–461. 1963.—A second transducing bacteriophage, designated SP-15, was isolated from the same soil-sample culture filtrate that supplied the Bacillus subtilis transducing phage, SP-10, reported earlier from this laboratory. SP-10 and SP-15 differ serologically and in several other respects, but share the ability to propagate on B. subtilis W-23-Sr (streptomycin-resistant) and B. licheniformis ATCC 9945a, and to mediate general transduction in either species when propagated homologously. Attempts to transduce between the species have failed. SP-10 forms plaques readily on both W-23-Sr and 9945a; SP-15 forms minute plaques on W-23-Sr and has shown no evidence of any lytic activity on 9945a. Maximal recoveries of prototrophic colonies from mixtures of SP-10 with auxotrophs of either W-23-Sr or 9945a were obtained only when excess phage was neutralized by post-transduction treatment with specific phage antiserum. Such treatment was not necessary for maximal recovery of transductants effected by SP-15. Unlike SP-10, SP-15 propagated on W-23-Sr did not transduce B. subtilis 168 (indole−). SP-15 transduced B. licheniformis more efficiently than did SP-10. Neither phage was able to transduce B. licheniformis as efficiently as it transduced B. subtilis. The differing influences of multiplicity of infection were compared for the two phages in both species. PMID:14066421

Differences in Shiga toxin and phage production among stx2g-positive STEC strains

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Claudia Viviana Granobles Velandia

2012-06-01

Full Text Available Shigatoxigenic E. coli (STEC are characterized by the production of Shiga toxins (Stx encoded by temperate bacteriophages. Stx production is linked to the induction of the phage lytic cycle. Several stx variants have been described and differentially associated with the risk of developing severe illness.The variant named stx2g was first identified in a STEC strain isolated from the faeces of healthy cattle. Analysis of stx2g-positive strains isolated from humans, animals and environmental sources have shown that they have a close relationship. In this study, stx2g-positive STEC isolated from cattle were analyzed for phage and Stx production, with the aim to relate the results to differences observed in cytotoxicity.The presence of inducible phages was assessed by analyzing the bacterial growth/lysis curves and also by plaque assay. Bacterial growth curves in the absence of induction were similar for all isolates, however, notably differed among induced cultures. The two strains that clearly evidenced bacteriolysis under this condition also showed higher phage titers in plaque assays. However, only the phage plaques produced by one of these strains (FB 62 hybridized with a stx2-probe. Furthermore, the production of Stx was evaluated by EIA and Western immunoblotting in overnight supernatants. By EIA, we detected Stx only in supernatants of FB 62, with a higher signal with induced than in uninduced cultures. By immunoblotting, Stx2 could be detected after induction in all stx2g-positive isolates, but with lower amounts of Stx2B subunit in those supernatants where phages could not be detected.Taking into account all the results, several differences could be found among stx2g-positive strains. The strain with the highest cytotoxic titer showed higher levels of stx2-phages and toxin production by EIA, and the opposite was observed for strains that previously showed low cytotoxic titers, confirming that in stx2g-positive strains Stx production is

Interaction between the genomes of Lactococcus lactis and phages of the P335 species

Science.gov (United States)

Kelly, William J.; Altermann, Eric; Lambie, Suzanne C.; Leahy, Sinead C.

2013-01-01

Phages of the P335 species infect Lactococcus lactis and have been particularly studied because of their association with strains of L. lactis subsp. cremoris used as dairy starter cultures. Unlike other lactococcal phages, those of the P335 species may have a temperate or lytic lifestyle, and are believed to originate from the starter cultures themselves. We have sequenced the genome of L. lactis subsp. cremoris KW2 isolated from fermented corn and found that it contains an integrated P335 species prophage. This 41 kb prophage (Φ KW2) has a mosaic structure with functional modules that are highly similar to several other phages of the P335 species associated with dairy starter cultures. Comparison of the genomes of 26 phages of the P335 species, with either a lytic or temperate lifestyle, shows that they can be divided into three groups and that the morphogenesis gene region is the most conserved. Analysis of these phage genomes in conjunction with the genomes of several L. lactis strains shows that prophage insertion is site specific and occurs at seven different chromosomal locations. Exactly how induced or lytic phages of the P335 species interact with carbohydrate cell surface receptors in the host cell envelope remains to be determined. Genes for the biosynthesis of a variable cell surface polysaccharide and for lipoteichoic acids (LTAs) are found in L. lactis and are the main candidates for phage receptors, as the genes for other cell surface carbohydrates have been lost from dairy starter strains. Overall, phages of the P335 species appear to have had only a minor role in the adaptation of L. lactis subsp. cremoris strains to the dairy environment, and instead they appear to be an integral part of the L. lactis chromosome. There remains a great deal to be discovered about their role, and their contribution to the evolution of the bacterial genome. PMID:24009606

The complete genome sequence and proteomics of Yersinia pestis phage Yep-phi.

Science.gov (United States)

Zhao, Xiangna; Wu, Weili; Qi, Zhizhen; Cui, Yujun; Yan, Yanfeng; Guo, Zhaobiao; Wang, Zuyun; Wang, Hu; Deng, Haijun; Xue, Yan; Chen, Weijun; Wang, Xiaoyi; Yang, Ruifu

2011-01-01

Yep-phi, a lytic phage of Yersinia pestis, was isolated in China and is routinely used as a diagnostic phage for the identification of the plague pathogen. Yep-phi has an isometric hexagonal head containing dsDNA and a short non-contractile conical tail. In this study, we sequenced the Yep-phi genome (GenBank accession no. HQ333270) and performed proteomics analysis. The genome consists of 38 ,616 bp of DNA, including direct terminal repeats of 222 bp, and is predicted to contain 45 ORFs. Most structural proteins were identified by proteomics analysis. Compared with the three available genome sequences of lytic phages for Y. pestis, the phages could be divided into two subgroups. Yep-phi displays marked homology to the bacteriophages Berlin (GenBank accession no. AM183667) and Yepe2 (GenBank accession no. EU734170), and these comprise one subgroup. The other subgroup is represented by bacteriophage ΦA1122 (GenBank accession no. AY247822). Potential recombination was detected among the Yep-phi subgroup.

Analytical Core Mass Function (CMF) from Filaments: Under Which Circumstances Can Filament Fragmentation Reproduce the CMF?

Energy Technology Data Exchange (ETDEWEB)

Lee, Yueh-Ning; Hennebelle, Patrick [IRFU, CEA, Université Paris-Saclay, F-91191 Gif-sur-Yvette (France); Chabrier, Gilles, E-mail: yueh-ning.lee@cea.fr [École normale supérieure de Lyon, CRAL, UMR CNRS 5574, Université de Lyon, F-69364 Lyon Cedex 07 (France)

2017-10-01

Observations suggest that star formation in filamentary molecular clouds occurs in a two-step process, with the formation of filaments preceding that of prestellar cores and stars. Here, we apply the gravoturbulent fragmentation theory of Hennebelle and Chabrier to a filamentary environment, taking into account magnetic support. We discuss the induced geometrical effect on the cores, with a transition from 3D geometry at small scales to 1D at large ones. The model predicts the fragmentation behavior of a filament for a given mass per unit length (MpL) and level of magnetization. This core mass function (CMF) for individual filaments is then convolved with the distribution of filaments to obtain the final system CMF. The model yields two major results. (i) The filamentary geometry naturally induces a hierarchical fragmentation process, first into groups of cores, separated by a length equal to a few filament Jeans lengths, i.e., a few times the filament width. These groups then fragment into individual cores. (ii) Non-magnetized filaments with high MpL are found to fragment excessively, at odds with observations. This is resolved by taking into account the magnetic field (treated simply as additional pressure support). The present theory suggests two complementary modes of star formation: although small (spherical or filamentary) structures will collapse directly into prestellar cores, according to the standard Hennebelle–Chabrier theory, the large (filamentary) ones, the dominant population according to observations, will follow the aforedescribed two-step process.

Automatic Detect and Trace of Solar Filaments

Science.gov (United States)

Fang, Cheng; Chen, P. F.; Tang, Yu-hua; Hao, Qi; Guo, Yang

We developed a series of methods to automatically detect and trace solar filaments in solar Hα images. The programs are able to not only recognize filaments and determine their properties, such as the position, the area and other relevant parameters, but also to trace the daily evolution of the filaments. For solar full disk Hα images, the method consists of three parts: first, preprocessing is applied to correct the original images; second, the Canny edge-detection method is used to detect the filaments; third, filament properties are recognized through the morphological operators. For each Hα filament and its barb features, we introduced the unweighted undirected graph concept and adopted Dijkstra shortest-path algorithm to recognize the filament spine; then, using polarity inversion line shift method for measuring the polarities in both sides of the filament to determine the filament axis chirality; finally, employing connected components labeling method to identify the barbs and calculating the angle between each barb and spine to indicate the barb chirality. Our algorithms are applied to the observations from varied observatories, including the Optical & Near Infrared Solar Eruption Tracer (ONSET) in Nanjing University, Mauna Loa Solar Observatory (MLSO) and Big Bear Solar Observatory (BBSO). The programs are demonstrated to be effective and efficient. We used our method to automatically process and analyze 3470 images obtained by MLSO from January 1998 to December 2009, and a butterfly diagram of filaments is obtained. It shows that the latitudinal migration of solar filaments has three trends in the Solar Cycle 23: The drift velocity was fast from 1998 to the solar maximum; after the solar maximum, it became relatively slow and after 2006, the migration became divergent, signifying the solar minimum. About 60% filaments with the latitudes larger than 50 degree migrate towards the Polar Regions with relatively high velocities, and the latitudinal migrating

High-resolution Observations of Sympathetic Filament Eruptions by NVST

Energy Technology Data Exchange (ETDEWEB)

Li, Shangwei; Su, Yingna; Zhou, Tuanhui; Ji, Haisheng [Key Laboratory for Dark Matter and Space Science, Purple Mountain Observatory, CAS, Nanjing 210008 (China); Van Ballegooijen, Adriaan [5001 Riverwood Avenue, Sarasota, FL 34231 (United States); Sun, Xudong, E-mail: ynsu@pmo.ac.cn [W. W. Hansen Experimental Physics Laboratory, Stanford University, Stanford, CA 94305 (United States)

2017-07-20

We investigate two sympathetic filament eruptions observed by the New Vacuum Solar Telescope on 2015 October 15. The full picture of the eruptions is obtained from the corresponding Solar Dynamics Observatory ( SDO )/Atmospheric Imaging Assembly (AIA) observations. The two filaments start from active region NOAA 12434 in the north and end in one large quiescent filament channel in the south. The left filament erupts first, followed by the right filament eruption about 10 minutes later. Clear twist structure and rotating motion are observed in both filaments during the eruption. Both eruptions failed, since the filaments first rise up, then flow toward the south and merge into the southern large quiescent filament. We also observe repeated activations of mini filaments below the right filament after its eruption. Using magnetic field models constructed based on SDO /HMI magnetograms via the flux rope insertion method, we find that the left filament eruption is likely to be triggered by kink instability, while the weakening of overlying magnetic fields due to magnetic reconnection at an X-point between the two filament systems might play an important role in the onset of the right filament eruption.

Semi-Solid and Solid Dosage Forms for the Delivery of Phage Therapy to Epithelia

Science.gov (United States)

Petrovski, Steve; Chan, Hiu Tat; Angove, Michael J.; Tucci, Joseph

2018-01-01

The delivery of phages to epithelial surfaces for therapeutic outcomes is a realistic proposal, and indeed one which is being currently tested in clinical trials. This paper reviews some of the known research on formulation of phages into semi-solid dosage forms such as creams, ointments and pastes, as well as solid dosage forms such as troches (or lozenges and pastilles) and suppositories/pessaries, for delivery to the epithelia. The efficacy and stability of these phage formulations is discussed, with a focus on selection of optimal semi-solid bases for phage delivery. Issues such as the need for standardisation of techniques for formulation as well as for assessment of efficacy are highlighted. These are important when trying to compare results from a range of experiments and across different delivery bases. PMID:29495355

Radiosensitivity of the induction of early enzymes by. gamma. -irradiated T7-phages

Energy Technology Data Exchange (ETDEWEB)

Bopp, E

1975-01-01

The radiosensitivity of the ability of the bacteriophage T7 to produce polymerase and lysozyme during its reproduction cycle is investigated. B-cells of Escherichia coli were infected with /sup 60/Co-..gamma..-irradiated T7 phages. From the extracts of the cells opened by ultrasonic waves, the amount of enzymes produced is determined with the aid of special enzyme tests. The fraction of inactivated phages able to produce RNA polymerase is higher than the fraction with intact DNA double strands and higher than the fraction able to inject DNA. The lowest fraction is that of inactivated phages producing lysozyme.

Bacteria between protists and phages: from antipredation strategies to the evolution of pathogenicity.

Science.gov (United States)

Brüssow, Harald

2007-08-01

Bacteriophages and protists are major causes of bacterial mortality. Genomics suggests that phages evolved well before eukaryotic protists. Bacteria were thus initially only confronted with phage predators. When protists evolved, bacteria were caught between two types of predators. One successful antigrazing strategy of bacteria was the elaboration of toxins that would kill the grazer. The released cell content would feed bystander bacteria. I suggest here that, to fight grazing protists, bacteria teamed up with those phage predators that concluded at least a temporary truce with them in the form of lysogeny. Lysogeny was perhaps initially a resource management strategy of phages that could not maintain infection chains. Subsequently, lysogeny might have evolved into a bacterium-prophage coalition attacking protists, which became a food source for them. When protists evolved into multicellular animals, the lysogenic bacteria tracked their evolving food source. This hypothesis could explain why a frequent scheme of bacterial pathogenicity is the survival in phagocytes, why a significant fraction of bacterial pathogens have prophage-encoded virulence genes, and why some virulence factors of animal pathogens are active against unicellular eukaryotes. Bacterial pathogenicity might thus be one playing option of the stone-scissor-paper game played between phages-bacteria-protists, with humans getting into the crossfire.

Advances in phage display technology for drug discovery.

Science.gov (United States)

Omidfar, Kobra; Daneshpour, Maryam

2015-06-01

Over the past decade, several library-based methods have been developed to discover ligands with strong binding affinities for their targets. These methods mimic the natural evolution for screening and identifying ligand-target interactions with specific functional properties. Phage display technology is a well-established method that has been applied to many technological challenges including novel drug discovery. This review describes the recent advances in the use of phage display technology for discovering novel bioactive compounds. Furthermore, it discusses the application of this technology to produce proteins and peptides as well as minimize the use of antibodies, such as antigen-binding fragment, single-chain fragment variable or single-domain antibody fragments like VHHs. Advances in screening, manufacturing and humanization technologies demonstrate that phage display derived products can play a significant role in the diagnosis and treatment of disease. The effects of this technology are inevitable in the development pipeline for bringing therapeutics into the market, and this number is expected to rise significantly in the future as new advances continue to take place in display methods. Furthermore, a widespread application of this methodology is predicted in different medical technological areas, including biosensing, monitoring, molecular imaging, gene therapy, vaccine development and nanotechnology.

Striation and convection in penumbral filaments

Science.gov (United States)

Spruit, H. C.; Scharmer, G. B.; Löfdahl, M. G.

2010-10-01

Observations with the 1-m Swedish Solar Telescope of the flows seen in penumbral filaments are presented. Time sequences of bright filaments show overturning motions strikingly similar to those seen along the walls of small isolated structures in the active regions. The filaments show outward propagating striations with inclination angles suggesting that they are aligned with the local magnetic field. We interpret it as the equivalent of the striations seen in the walls of small isolated magnetic structures. Their origin is then a corrugation of the boundary between an overturning convective flow inside the filament and the magnetic field wrapping around it. The outward propagation is a combination of a pattern motion due to the downflow observed along the sides of bright filaments, and the Evershed flow. The observed short wavelength of the striation argues against the existence of a dynamically significant horizontal field inside the bright filaments. Its intensity contrast is explained by the same physical effect that causes the dark cores of filaments, light bridges and “canals”. In this way striation represents an important clue to the physics of penumbral structure and its relation with other magnetic structures on the solar surface. We put this in perspective with results from the recent 3-D radiative hydrodynamic simulations. 4 movies are only available in electronic form at http://www.aanda.org

Emergence of new Salmonella Enteritidis phage types in Europe? Surveillance of infections in returning travellers

Directory of Open Access Journals (Sweden)

Andersson Yvonne

2004-09-01

Full Text Available Abstract Background Among human Salmonella Enteritidis infections, phage type 4 has been the dominant phage type in most countries in Western Europe during the last years. This is reflected in Salmonella infections among Swedish travellers returning from abroad. However, there are differences in phage type distribution between the countries, and this has also changed over time. Methods We used data from the Swedish infectious disease register and the national reference laboratory to describe phage type distribution of Salmonella Enteritidis infections in Swedish travellers from 1997 to 2002, and have compared this with national studies conducted in the countries visited. Results Infections among Swedish travellers correlate well with national studies conducted in the countries visited. In 2001 a change in phage type distribution in S. Enteritidis infections among Swedish travellers returning from some countries in southern Europe was observed, and a previously rare phage type (PT 14b became one of the most commonly diagnosed that year, continuing into 2002 and 2003. Conclusions Surveillance of infections among returning travellers can be helpful in detecting emerging infections and outbreaks in tourist destinations. The information needs to be communicated rapidly to all affected countries in order to expedite the implementation of appropriate investigations and preventive measures.

Phages of Lactobacillus casei/paracasei: response to environmental factors and interaction with collection and commercial strains.

Science.gov (United States)

Capra, M L; Quiberoni, A; Reinheimer, J

2006-02-01

To investigate the influence of several environmental factors on the viability and cell-adsorption for two Lactobacillus casei/paracasei bacteriophages (PL-1 and J-1). Both phages showed a remarkably high specificity of species, sharing similar host spectra. Two phages and four sensitive strains were used to conform five phage/strain systems. Each showed a particular behaviour (burst size: ranging from 32 to 160 PFU/infective centre; burst time: 120-240 min and latent time: 5-90 min). For both phages, the viability was not significantly affected from pH 4 to 11 (room temperature) and from pH 5 to 10 (37 degrees C). Adsorption rates were not influenced by calcium ions, but decreased after the thermal inactivation of cells. Adsorption rates were high between 0 and 50 degrees C with maximum values at 30 degrees C and pH 6. System PL-1/Lact. paracasei A showed noticeable differences in comparison with the others, being times required to reach 90% of adsorption of 4 h and lower than 45 min, respectively. The data obtained in this work demonstrated that environmental parameters can influence the viability and cell adsorption rates of Lact. casei/paracasei phages. The extent of this influence was phage dependent. This work contributes to the enlargement of the currently scarce knowledge of phages of probiotic bacteria.

Evolution of filament barbs.

Science.gov (United States)

Liu, R.; Xu, Y.; Wang, H.

We present a selected few cases in which the sense of chirality of filament barbs changed within periods as short as hours. We investigate in detail a quiescent filament on 2003 September 10 and 11. Of its four barbs displaying such changes, only one overlays a small polarity inversion line inside the EUV filament channel (EFC). No magnetic elements with magnitude above the noise level were detected at the endpoints of all barbs. In particular, a pair of barbs first approached toward, and then departed from, each other in Halpha , with the barb endpoints migrating as far as ˜ 10 arcsec. We conclude that the evolution of the barbs was driven by flux emergence and cancellation of small bipolar units at the EFC border.

Clustered Regularly Interspaced Short Palindromic Repeat-Dependent, Biofilm-Specific Death of Pseudomonas aeruginosa Mediated by Increased Expression of Phage-Related Genes.

Science.gov (United States)

Heussler, Gary E; Cady, Kyle C; Koeppen, Katja; Bhuju, Sabin; Stanton, Bruce A; O'Toole, George A

2015-05-12

-virus interplay but has also led to a major advancement in genetic engineering. Recently, increasing evidence suggested that bacteria can co-opt the CRISPR system for functions besides adaptive immunity to phage infection. This study examined one such alternative function, and this report describes the mechanism of type 1-F CRISPR-dependent loss of the biofilm and swarming in the medically relevant opportunistic pathogen Pseudomonas aeruginosa. Since both biofilm formation and swarming motility are important in the virulence of P. aeruginosa, a full understanding of how the CRISPR system can regulate such group behaviors is fundamental to developing new therapeutics. Copyright © 2015 Heussler et al.

Characterization and lytic activity of methicillin-resistant Staphylococcus aureus(MRSA phages isolated from NICU

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Golnar Rahimzadeh

2016-06-01

Full Text Available Background Methicillin-resistant Staphylococcus aureus (MRSA is a well-known pathogen that causes serious diseases in humans. As part of the efforts to control this pathogen, an isolated bacteriophage, Siphoviridae, which specifically targets Methicillin-resistant Staphylococcus aureus (MRSA, was characterized. Aims The objective of this study was to characterize of a virulent bacteriophage (Siphoviridae isolated from a NICU bathroom sink. Methods The MRSA strain was isolated from patient blood. The isolated strain was confirmed as MRSA using conventional methods. Phages were isolated from a NICU bathroom sink and activity was lytic as determined by spot test. Titer phage lysate was measured by the Double Layer Agar (DLA technique. The morphology was found with electron microscopy. The single-step growth curve was plotted. Results Electron microscopy showed the phage as a member of the family Siphoviridae, serogroup A and F. The isolated phage was capable of lytic activity against methicillin-resistant Staphylococcus aureus (MRSA strain as shown by spot test. By DLA, the titre of the phages was determined to be 10×108PFU/ml. The single-step growth curve showed that the latent period of the isolated bacteriophage was 30 min and the total number of viable progeny per infected host, burst size, was 2600 PFU/infected host. Conclusion In this study, two phages were isolated and characterized from a NICU bathroom sink, from the Siphoviridae family, which specifically targetsmethicillin-resistant Staphylococcus aureus (MRSA.

Development of anti-infectives using phage display: biological agents against bacteria, viruses, and parasites.

Science.gov (United States)

Huang, Johnny X; Bishop-Hurley, Sharon L; Cooper, Matthew A

2012-09-01

The vast majority of anti-infective therapeutics on the market or in development are small molecules; however, there is now a nascent pipeline of biological agents in development. Until recently, phage display technologies were used mainly to produce monoclonal antibodies (MAbs) targeted against cancer or inflammatory disease targets. Patent disputes impeded broad use of these methods and contributed to the dearth of candidates in the clinic during the 1990s. Today, however, phage display is recognized as a powerful tool for selecting novel peptides and antibodies that can bind to a wide range of antigens, ranging from whole cells to proteins and lipid targets. In this review, we highlight research that exploits phage display technology as a means of discovering novel therapeutics against infectious diseases, with a focus on antimicrobial peptides and antibodies in clinical or preclinical development. We discuss the different strategies and methods used to derive, select, and develop anti-infectives from phage display libraries and then highlight case studies of drug candidates in the process of development and commercialization. Advances in screening, manufacturing, and humanization technologies now mean that phage display can make a significant contribution in the fight against clinically important pathogens.

Potential effect of some environmental factors on the phage removal during wastewater treatment. Study in vitro

International Nuclear Information System (INIS)

Benhyahya, M.; Bohatier, J.; Laveran, H.; Ettayebi, M.; Senaud, J.

2000-01-01

Great quantities of enteric viruses and bacteriophages are included in wastewaters. They represent a contamination risk of natural water systems. But this viral burden is greatly reduced in the sewage treatment plants by the combined action of numerous environmental factors. To study water quality, some groups of bacteriophages as E. coli phages and Bacteroides fragilis phages have been proposed as model viruses. On an other hand, somatic and, in particular, F-specific coliphages have several morphological, structural and chemical composition ressemblances with the enteric viruses. Two different bacteriophages (øX-174 and MS2) were used as virus models in this in vitro study to evaluate the viral adsorption on suspended clay particles. Distilled sterile water was used as reactional medium to avoid the possible interactions with the considered substrates, the kaolinite (K) and the montmorillonite (M). Phage behaviour in the water and in the recommended diluent for phages, the saline peptone, was first compared. K and M suspensions were used at 300 mg/l for a contact time of 5, 30 and 60 min. In other series K and M suspensions were prepared at 600, 300 and 100 mg/l then used to determine the phage adsorption capacity in a fixed time 30 min. Results show that the phage titers for all samples were constant in the organic diluent. They were lower in the distilled sterile water and decrease with the time. Distilled water favours most likely the grouping of virions and leads aggregates formation. The adsorption of øX-174 and MS2 onto K or M particles was instantaneous and independent of the duration contact. The clay concentration had a slight significant influence on the phage adsorption rate. Using the same phages we have studied, in a second stage, the potential effect of the dissolved matters in a filtered polluted effluent, that of sunlight radiations and that of the protozoan Tetrahymena pyriformis on the phage removal. No soon significant phage inactivation was

Phage nanofibers induce vascularized osteogenesis in 3D printed bone scaffolds.

Science.gov (United States)

Wang, Jianglin; Yang, Mingying; Zhu, Ye; Wang, Lin; Tomsia, Antoni P; Mao, Chuanbin

2014-08-06

A virus-activated matrix is developed to overcome the challenge of forming vascularized bone tissue. It is generated by filling a 3D printed bioceramic scaffold with phage nanofibers displaying high-density RGD peptide. After it is seeded with mesenchymal stem cells (MSCs) and implanted into a bone defect, the phage nanofibers induce osteogenesis and angiogenesis by activating endothelialization and osteogenic differentiation of MSCs. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Damages induced in lambda phage DNA by enzyme-generated triplet acetone

International Nuclear Information System (INIS)

Menck, C.F.; Cabral Neto, J.B.; Gomes, R.A.; Faljoni-Alario, A.

1985-01-01

Exposure of lambda phage to triplet acetone, generated during the aerobic oxidation of isobutanal by peroxidase, leads to genome lesions. The majority of these lesions are detected as DNA single-strand breaks only in alkaline conditions, so true breaks were not observed. Also, no sites sensitive to UV-endonuclease from Micrococcus luteus were found in DNA from treated phage. The participation of triplet acetone in the generation of such DNA damage is discussed. (Author) [pt

Identification of antigenic regions on VP2 of African horsesickness virus serotype 3 by using phage-displayed epitope libraries.

Science.gov (United States)

Bentley, L; Fehrsen, J; Jordaan, F; Huismans, H; du Plessis, D H

2000-04-01

VP2 is an outer capsid protein of African horsesickness virus (AHSV) and is recognized by serotype-discriminatory neutralizing antibodies. With the objective of locating its antigenic regions, a filamentous phage library was constructed that displayed peptides derived from the fragmentation of a cDNA copy of the gene encoding VP2. Peptides ranging in size from approximately 30 to 100 amino acids were fused with pIII, the attachment protein of the display vector, fUSE2. To ensure maximum diversity, the final library consisted of three sub-libraries. The first utilized enzymatically fragmented DNA encoding only the VP2 gene, the second included plasmid sequences, while the third included a PCR step designed to allow different peptide-encoding sequences to recombine before ligation into the vector. The resulting composite library was subjected to immunoaffinity selection with AHSV-specific polyclonal chicken IgY, polyclonal horse immunoglobulins and a monoclonal antibody (MAb) known to neutralize AHSV. Antigenic peptides were located by sequencing the DNA of phages bound by the antibodies. Most antigenic determinants capable of being mapped by this method were located in the N-terminal half of VP2. Important binding areas were mapped with high resolution by identifying the minimum overlapping areas of the selected peptides. The MAb was also used to screen a random 17-mer epitope library. Sequences that may be part of a discontinuous neutralization epitope were identified. The amino acid sequences of the antigenic regions on VP2 of serotype 3 were compared with corresponding regions on three other serotypes, revealing regions with the potential to discriminate AHSV serotypes serologically.

Development of the striation and filament form of the electrothermal instability

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Yu, Edmund; Awe, T. J.; Yelton, W. G.; McKenzie, B. B.; Peterson, K. J.; Bauer, B. S.; Hutchinson, T. M.; Fuelling, S.; Yates, K. C.; Shipley, G.

2017-10-01

Magnetically imploded liners have broad application to ICF, dynamic material property studies, and flux compression. An important consideration in liner performance is the electrothermal instability (ETI), an Ohmic heating instability that manifests in 2 ways: assuming vertical current flow, ETI forms hot, horizontal bands (striations) in metals, and vertical filaments in plasmas. Striations are especially relevant in that they can develop into density perturbations, which then couple to the dangerous magneto Rayleigh-Taylor (MRT) instability during liner acceleration. Recent visible emission images of Ohmically heated rods show evidence of both the striation and filament form of ETI, suggesting several questions: (1) can simulation qualitatively reproduce the data? (2) If so, what seeds the striation ETI, and how does it transition to filaments? (3) Does the striation develop into a strong density perturbation, important for MRT? In this work, we use analytic theory and 3D MHD simulation to study how isolated resistive inclusions, embedded in a perfectly smooth rod and communicating through current redistribution, can be used to address the above questions. Sandia National Laboratories is a multimission laboratory managed and operated by National Technology and Engineering Solutions of Sandia LLC, a wholly owned subsidiary of Honeywell International Inc. for the U.S. DOE NNSA under contract DE-NA0003525.

Filamentation of Campylobacter in broth cultures

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Nacheervan M Ghaffar

2015-06-01

Full Text Available The transition from rod to filamentous cell morphology has been identified as a response to stressful conditions in many bacterial species and has been ascribed to confer certain survival advantages. Filamentation of Campylobacter jejuni was demonstrated to occur spontaneously on entry in to stationary phase distinguishing it from many other bacteria where a reduction in size is more common. The aim of this study was to investigate the cues that give rise to filamentation of C. jejuni and C. coli and gain insights into the process. Using minimal medium, augmentation of filamentation occurred and it was observed that this morphological change was wide spread amongst C. jejuni strains tested but was not universal in C. coli strains. Filamentation did not appear to be due to release of diffusible molecules, toxic metabolites, or be in response to oxidative stress in the medium. Separated filaments exhibited greater intracellular ATP contents (2.66 to 17.4 fg than spiral forms (0.99 to 1.7 fg and showed enhanced survival in water at 4oC and 37oC compared to spiral cells. These observations support the conclusion that the filaments are adapted to survive extra-intestinal environments. Differences in cell morphology and physiology need to be considered in the context of the design of experimental studies and the methods adopted for the isolation of campylobacters from food, clinical and environmental sources.

Effectiveness of the lactococcal abortive infection systems AbiA, AbiE, AbiF and AbiG against P335 type phages.

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Tangney, Mark; Fitzgerald, Gerald F

2002-04-23

Four lactococcal abortive infection mechanisms were introduced into strains which were sensitive hosts for P335 type phages and plaque assay experiments performed to assess their effect on five lactococcal bacteriophages from this family. Results indicate that AbiA inhibits all five P335 phages tested, while AbiG affects phiP335 itself and phiQ30 but not the other P335 species phages. AbiA was shown to retard phage Q30 DNA replication as previously reported for other phages. It was also demonstrated that AbiG, previously shown to act at a point after DNA replication in the cases of c2 type and 936 type phages, acts at the level of, or prior to phage Q30 DNA replication. AbiE and AbiF had no effect on the P335 type phages examined.

Transcriptome Analysis of a Bloom-Forming Cyanobacterium Microcystis aeruginosa during Ma-LMM01 Phage Infection

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Daichi Morimoto

2018-01-01

Full Text Available Microcystis aeruginosa forms massive blooms in eutrophic freshwaters, where it is constantly exposed to lytic cyanophages. Unlike other marine cyanobacteria, M. aeruginosa possess remarkably abundant and diverse potential antiviral defense genes. Interestingly, T4-like cyanophage Ma-LMM01, which is the sole cultured lytic cyanophage infecting M. aeruginosa, lacks the host-derived genes involved in maintaining host photosynthesis and directing host metabolism that are abundant in other marine cyanophages. Based on genomic comparisons with closely related cyanobacteria and their phages, Ma-LMM01 is predicted to employ a novel infection program that differs from that of other marine cyanophages. Here, we used RNA-seq technology and in silico analysis to examine transcriptional dynamics during Ma-LMM01 infection to reveal host transcriptional responses to phage infection, and to elucidate the infection program used by Ma-LMM01 to avoid the highly abundant host defense systems. Phage-derived reads increased only slightly at 1 h post-infection, but significantly increased from 16% of total cellular reads at 3 h post-infection to 33% of all reads by 6 h post-infection. Strikingly, almost none of the host genes (0.17% showed a significant change in expression during infection. However, like other lytic dsDNA phages, including marine cyanophages, phage gene dynamics revealed three expression classes: early (host-takeover, middle (replication, and late (virion morphogenesis. The early genes were concentrated in a single ∼5.8-kb window spanning 10 open reading frames (gp054–gp063 on the phage genome. None of the early genes showed homology to the early genes of other T4-like phages, including known marine cyanophages. Bacterial RNA polymerase (σ70 recognition sequences were also found in the upstream region of middle and late genes, whereas phage-specific motifs were not found. Our findings suggest that unlike other known T4-like phages, Ma-LMM01

Plasma Brightenings in a Failed Solar Filament Eruption

Energy Technology Data Exchange (ETDEWEB)

Li, Y.; Ding, M. D., E-mail: yingli@nju.edu.cn [School of Astronomy and Space Science, Nanjing University, Nanjing 210023 (China)

2017-03-20

Failed filament eruptions are solar eruptions that are not associated with coronal mass ejections. In a failed filament eruption, the filament materials usually show some ascending and falling motions as well as generating bright EUV emissions. Here we report a failed filament eruption (SOL2016-07-22) that occurred in a quiet-Sun region observed by the Atmospheric Imaging Assembly on board the Solar Dynamics Observatory . In this event, the filament spreads out but gets confined by the surrounding magnetic field. When interacting with the ambient magnetic field, the filament material brightens up and flows along the magnetic field lines through the corona to the chromosphere. We find that some materials slide down along the lifting magnetic structure containing the filament and impact the chromosphere, and through kinetic energy dissipation, cause two ribbon-like brightenings in a wide temperature range. There is evidence suggesting that magnetic reconnection occurs between the filament magnetic structure and the surrounding magnetic fields where filament plasma is heated to coronal temperatures. In addition, thread-like brightenings show up on top of the erupting magnetic fields at low temperatures, which might be produced by an energy imbalance from a fast drop of radiative cooling due to plasma rarefaction. Thus, this single event of a failed filament eruption shows the existence of a variety of plasma brightenings that may be caused by completely different heating mechanisms.

Stability of Staphylococcus aureus phage ISP after freeze-drying (lyophilization.

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Maia Merabishvili

Full Text Available Staphylococcus aureus phage ISP was lyophilized, using an Amsco-Finn Aqua GT4 freeze dryer, in the presence of six different stabilizers at different concentrations. Stability of the lyophilized phage at 4 °C was monitored up to 37 months and compared to stability in Luria Bertani broth and physiological saline at 4 °C. Sucrose and trehalose were shown to be the best stabilizing additives, causing a decrease of only 1 log immediately after the lyophilization procedure and showing high stability during a 27 month storage period.

Construction and Selection of Affilin® Phage Display Libraries.

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Settele, Florian; Zwarg, Madlen; Fiedler, Sebastian; Koscheinz, Daniel; Bosse-Doenecke, Eva

2018-01-01

Affilin ® molecules represent a new class of so-called scaffold proteins. The concept of scaffold proteins is to use stable and versatile protein structures which can be endowed with de novo binding properties and specificities by introducing mutations in surface exposed amino acid residues. Complex variations and combinations are generated by genetic methods of randomization resulting in large cDNA libraries. The selection for candidates binding to a desired target can be executed by display methods, especially the very robust and flexible phage display. Here, we describe the construction of ubiquitin based Affilin ® phage display libraries and their use in biopanning experiments for the identification of novel protein ligands.

Graphene-based filament material for thermal ionization

Energy Technology Data Exchange (ETDEWEB)

Hewitt, J. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Shick, C. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Siegfried, M. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

2017-09-19

The use of graphene oxide materials for thermal ionization mass spectrometry analysis of plutonium and uranium has been investigated. Filament made from graphene oxide slurries have been 3-D printed. A method for attaching these filaments to commercial thermal ionization post assemblies has been devised. Resistive heating of the graphene based filaments under high vacuum showed stable operation in excess of 4 hours. Plutonium ion production has been observed in an initial set of filaments spiked with the Pu 128 Certified Reference Material.

Complete nucleotide sequence of Bacillus subtilis (natto) bacteriophage PM1, a phage associated with disruption of food production.

Science.gov (United States)

Umene, Kenichi; Shiraishi, Atsushi

2013-06-01

"Natto", considered a traditional food, is made by fermenting boiled soybeans with Bacillus subtilis (natto), which is a natto-producing strain related to B. subtilis. The production of natto is disrupted by phage infections of B. subtilis (natto); hence, it is necessary to control phage infections. PM1, a phage of B. subtilis (natto), was isolated during interrupted natto production in a factory. In a previous study, PM1 was classified morphologically into the family Siphoviridae, and its genome, comprising approximately 50 kbp of linear double-stranded DNA, was assumed to be circularly permuted. In the present study, the complete nucleotide sequence of the PM1 genomic DNA of 50,861 bp (41.3 %G+C) was determined, and 86 open reading frames (ORFs) were deduced. Forty-one ORFs of PM1 shared similarities with proteins deduced from the genome of phages reported so far. Twenty-three ORFs of PM1 were associated with functions related to the phage multiplication process of gene control, DNA replication/modification, DNA packaging, morphogenesis, and cell lysis. Bacillus subtilis (natto) produces a capsular polypeptide of glutamate with a γ-linkage (called poly-γ-glutamate), which appears to serve as a physical barrier to phage adsorption. One ORF of PM1 had similarity with a poly-γ-glutamate hydrolase, which is assumed to degrade the capsular barrier to allow phage progenies to infect encapsulated host cells. The genome analysis of PM1 revealed the characteristics of the phage that are consistent as Bacillus subtilis (natto)-infecting phage.

Characterization and Complete Genome Sequences of Three N4-Like Roseobacter Phages Isolated from the South China Sea.

Science.gov (United States)

Li, Baolian; Zhang, Si; Long, Lijuan; Huang, Sijun

2016-09-01

Three bacteriophages (RD-1410W1-01, RD-1410Ws-07, and DS-1410Ws-06) were isolated from the surface water of Sanya Bay, northern South China Sea, on two marine bacteria type strains of the Roseobacter lineage. These phages have an isometric head and a short tail, morphologically belonging to the Podoviridae family. Two of these phages can infect four of seven marine roseobacter strains tested and the other one can infect three of them, showing relatively broader host ranges compared to known N4-like roseophages. One-step growth curves showed that these phages have similar short latent periods (1-2 h) but highly variable burst sizes (27-341 pfu cell(-1)). Their complete genomes show high level of similarities to known N4-like roseophages in terms of genome size, G + C content, gene content, and arrangement. The morphological and genomic features of these phages indicate that they belong to the N4likevirus genus. Moreover, comparative genomic analysis based on 43 N4-like phages (10 roseobacter phages and 33 phages infecting other lineages of bacteria) revealed a core genome of 18 genes shared by all the 43 phages and 38 genes shared by all the ten roseophages. The 38 core genes of N4-like roseophages nearly make up 70 % of each genome in length. Phylogenetic analysis based on the concatenated core gene products showed that our phage isolates represent two new phyletic branches, suggesting the broad genetic diversity of marine N4-like roseophages remains.

Next-generation phage display: integrating and comparing available molecular tools to enable cost-effective high-throughput analysis.

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Emmanuel Dias-Neto

2009-12-01

Full Text Available Combinatorial phage display has been used in the last 20 years in the identification of protein-ligands and protein-protein interactions, uncovering relevant molecular recognition events. Rate-limiting steps of combinatorial phage display library selection are (i the counting of transducing units and (ii the sequencing of the encoded displayed ligands. Here, we adapted emerging genomic technologies to minimize such challenges.We gained efficiency by applying in tandem real-time PCR for rapid quantification to enable bacteria-free phage display library screening, and added phage DNA next-generation sequencing for large-scale ligand analysis, reporting a fully integrated set of high-throughput quantitative and analytical tools. The approach is far less labor-intensive and allows rigorous quantification; for medical applications, including selections in patients, it also represents an advance for quantitative distribution analysis and ligand identification of hundreds of thousands of targeted particles from patient-derived biopsy or autopsy in a longer timeframe post library administration. Additional advantages over current methods include increased sensitivity, less variability, enhanced linearity, scalability, and accuracy at much lower cost. Sequences obtained by qPhage plus pyrosequencing were similar to a dataset produced from conventional Sanger-sequenced transducing-units (TU, with no biases due to GC content, codon usage, and amino acid or peptide frequency. These tools allow phage display selection and ligand analysis at >1,000-fold faster rate, and reduce costs approximately 250-fold for generating 10(6 ligand sequences.Our analyses demonstrates that whereas this approach correlates with the traditional colony-counting, it is also capable of a much larger sampling, allowing a faster, less expensive, more accurate and consistent analysis of phage enrichment. Overall, qPhage plus pyrosequencing is superior to TU-counting plus Sanger

Footpoint detection and mass-motion in chromospheric filaments

Science.gov (United States)

V, Aparna; Hardersen, P. S.; Martin, S. F.

2013-07-01

A quiescent region on the Sun containing three filaments is used to study the properties of mass motion. This study determines if the footpoints or end-points of the filaments are the locations from where mass gets injected into the filaments. Several hypotheses have been put forth in the past to determine how a filament acquires mass. Trapping of coronal mass in the filament channel due to condensation (Martin, 1996) and injection of mass into the filaments during magnetic reconnection (Priest, et al., 1995) are some of the speculations. This study looks for indications for injection of mass via chromospheric footpoints. The data consists of blue (Hα-0.5 Å) and red (Hα+0.5 Å) wing high resolution Hα images of the W29N37 region of the Sun taken on Oct 30, 2010, from 1200 - 1600 UT. The Dutch Open Telescope was used to obtain the data. The images are aligned and animated to see Doppler motion in the fibrils. Smaller fibrils merge to form longer ones; barbs appear and disappear in one of the long filaments and is seen moving along the length of the filament. A region with no typical filament-like absorption feature is observed to be continuously receiving mass. Fibrils appear to be converging from opposite sides along what appears to be a neutral line; mass motion is seen in these fibrils as well. An eruption occurs in a region of fibrils lumped together at the end of the first hour (1300 UT) followed by plage brightening at 1430 UT near one of the filament regions. Helioviewer (Panasenco, et al., 2011) is used for aligning the images; GIMP is used for precision alignment and animation. Each frame in the sequence is studied carefully to note changes in the filament regions. The footpoints of the filaments are determined by the changes observed in the position of the filament ‘legs’ in each frame. Variations in the magnetic polarity corresponding to changes observed in the chromosphere are analyzed using HMI magnetograms. Bright and dark points on the

Thick Filament Protein Network, Functions, and Disease Association.

Science.gov (United States)

Wang, Li; Geist, Janelle; Grogan, Alyssa; Hu, Li-Yen R; Kontrogianni-Konstantopoulos, Aikaterini

2018-03-13

Sarcomeres consist of highly ordered arrays of thick myosin and thin actin filaments along with accessory proteins. Thick filaments occupy the center of sarcomeres where they partially overlap with thin filaments. The sliding of thick filaments past thin filaments is a highly regulated process that occurs in an ATP-dependent manner driving muscle contraction. In addition to myosin that makes up the backbone of the thick filament, four other proteins which are intimately bound to the thick filament, myosin binding protein-C, titin, myomesin, and obscurin play important structural and regulatory roles. Consistent with this, mutations in the respective genes have been associated with idiopathic and congenital forms of skeletal and cardiac myopathies. In this review, we aim to summarize our current knowledge on the molecular structure, subcellular localization, interacting partners, function, modulation via posttranslational modifications, and disease involvement of these five major proteins that comprise the thick filament of striated muscle cells. © 2018 American Physiological Society. Compr Physiol 8:631-709, 2018. Copyright © 2018 American Physiological Society. All rights reserved.

Probing ADAMTS13 substrate specificity using phage display.

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Karl C Desch

Full Text Available Von Willebrand factor (VWF is a large, multimeric protein that regulates hemostasis by tethering platelets to the subendothelial matrix at sites of vascular damage. The procoagulant activity of plasma VWF correlates with the length of VWF multimers, which is proteolytically controlled by the metalloprotease ADAMTS13. To probe ADAMTS13 substrate specificity, we created phage display libraries containing randomly mutated residues of a minimal ADAMTS13 substrate fragment of VWF, termed VWF73. The libraries were screened for phage particles displaying VWF73 mutant peptides that were resistant to proteolysis by ADAMTS13. These peptides exhibited the greatest mutation frequency near the ADAMTS13 scissile residues. Kinetic assays using mutant and wild-type substrates demonstrated excellent agreement between rates of cleavage for mutant phage particles and the corresponding mutant peptides. Cleavage resistance of selected mutations was tested in vivo using hydrodynamic injection of corresponding full-length expression plasmids into VWF-deficient mice. These studies confirmed the resistance to cleavage resulting from select amino acid substitutions and uncovered evidence of alternate cleavage sites and recognition by other proteases in the circulation of ADAMTS13 deficient mice. Taken together, these studies demonstrate the key role of specific amino acids residues including P3-P2' and P11', for substrate specificity and emphasize the importance in flowing blood of other ADAMTS13-VWF exosite interactions outside of VWF73.

Molecular and chemical engineering of bacteriophages for potential medical applications.

Science.gov (United States)

Hodyra, Katarzyna; Dąbrowska, Krystyna

2015-04-01

Recent progress in molecular engineering has contributed to the great progress of medicine. However, there are still difficult problems constituting a challenge for molecular biology and biotechnology, e.g. new generation of anticancer agents, alternative biosensors or vaccines. As a biotechnological tool, bacteriophages (phages) offer a promising alternative to traditional approaches. They can be applied as anticancer agents, novel platforms in vaccine design, or as target carriers in drug discovery. Phages also offer solutions for modern cell imaging, biosensor construction or food pathogen detection. Here we present a review of bacteriophage research as a dynamically developing field with promising prospects for further development of medicine and biotechnology.

Filament shape versus coronal potential magnetic field structure

Science.gov (United States)

Filippov, B.

2016-01-01

Solar filament shape in projection on disc depends on the structure of the coronal magnetic field. We calculate the position of polarity inversion lines (PILs) of coronal potential magnetic field at different heights above the photosphere, which compose the magnetic neutral surface, and compare with them the distribution of the filament material in Hα chromospheric images. We found that the most of the filament material is enclosed between two PILs, one at a lower height close to the chromosphere and one at a higher level, which can be considered as a height of the filament spine. Observations of the same filament on the limb by the Solar Terrestrial Relations Observatory spacecraft confirm that the height of the spine is really very close to the value obtained from the PIL and filament border matching. Such matching can be used for filament height estimations in on-disc observations. Filament barbs are housed within protruding sections of the low-level PIL. On the base of simple model, we show that the similarity of the neutral surfaces in potential and non-potential fields with the same sub-photospheric sources is the reason for the found tendency for the filament material to gather near the potential-field neutral surface.

Modular architecture of the T4 phage superfamily: A conserved core genome and a plastic periphery

International Nuclear Information System (INIS)

Comeau, Andre M.; Bertrand, Claire; Letarov, Andrei; Tetart, Francoise; Krisch, H.M.

2007-01-01

Among the most numerous objects in the biosphere, phages show enormous diversity in morphology and genetic content. We have sequenced 7 T4-like phages and compared their genome architecture. All seven phages share a core genome with T4 that is interrupted by several hyperplastic regions (HPRs) where most of their divergence occurs. The core primarily includes homologues of essential T4 genes, such as the virion structure and DNA replication genes. In contrast, the HPRs contain mostly novel genes of unknown function and origin. A few of the HPR genes that can be assigned putative functions, such as a series of novel Internal Proteins, are implicated in phage adaptation to the host. Thus, the T4-like genome appears to be partitioned into discrete segments that fulfil different functions and behave differently in evolution. Such partitioning may be critical for these large and complex phages to maintain their flexibility, while simultaneously allowing them to conserve their highly successful virion design and mode of replication

On the lack of host-cell reactivation of UV-irradiated phage T5

International Nuclear Information System (INIS)

Chiang, T.; Harm, W.

1976-01-01

Previously reported experiments have shown that host-cell reactivation (HCR) of UV-irradiated phage T1 in excision-repair proficient Escherichia coli cells is inhibited by superinfection with phage T5. Theoretical considerations have led to predictions concerning the dependence of repair inhibition on the multiplicity of superinfecting T5 phage and on the UV fluence to which they were exposed. These predictions have been supported by experimental results described in this paper. The fluence dependence permitted calculation of the relative UV sensitivity of the gene function responsible for repair inhibition; it was found to be about 2.3% that of the plaque-forming ability of phage T5. The T5-inhibitable step in excision repair occurs early in the infective cycle of T1. Furthermore, experiments involving the presence of 400 μg/ml chloramphenicol showed that HCR inhibition of T1 is caused by a protein produced after the FST segment of T5 (i.e. the first 8% of the T5 genome) has entered the host cell. A previously described minor T1 recovery process, occuring in both excision-repair-proficient and -deficient host-cells, is inhibited by T5 infection due to a different substance, which is most likely associated with the 'second-step-transfer' region of T5 DNA (involving the remainder of the genome). Superinfection with T4v 1 phage resulted in HCR inhibition of T1, resembling that observed after T5 superinfection. The discussion of these results suggests that inhibition of the bacterial excision repair system by T5 or T4 infection occurs at the level of UV-endonucleolytic incision, and that lack of HCR both in T-even phages and in T5 can be explained in the same manner

Fabrication of PLA Filaments and its Printable Performance

Science.gov (United States)

Liu, Wenjie; Zhou, Jianping; Ma, Yuming; Wang, Jie; Xu, Jie

2017-12-01

Fused deposition modeling (FDM) is a typical 3D printing technology and preparation of qualified filaments is the basis. In order to prepare polylactic acid (PLA) filaments suitable for personalized FDM 3D printing, this article investigated the effect of factors such as extrusion temperature and screw speed on the diameter, surface roughness and ultimate tensile stress of the obtained PLA filaments. The optimal process parameters for fabrication of qualified filaments were determined. Further, the printable performance of the obtained PLA filaments for 3D objects was preliminarily explored.

Terahertz waves radiated from two noncollinear femtosecond plasma filaments

Energy Technology Data Exchange (ETDEWEB)

Du, Hai-Wei; Hoshina, Hiromichi; Otani, Chiko, E-mail: otani@riken.jp [Terahertz Sensing and Imaging Research Team, RIKEN Center for Advanced Photonics, RIKEN, Sendai, Miyagi 980-0845 (Japan); Midorikawa, Katsumi [Attosecond Science Research Team, RIKEN Center for Advanced Photonics, RIKEN, Wako, Saitama 351-0198 (Japan)

2015-11-23

Terahertz (THz) waves radiated from two noncollinear femtosecond plasma filaments with a crossing angle of 25° are investigated. The irradiated THz waves from the crossing filaments show a small THz pulse after the main THz pulse, which was not observed in those from single-filament scheme. Since the position of the small THz pulse changes with the time-delay of two filaments, this phenomenon can be explained by a model in which the small THz pulse is from the second filament. The denser plasma in the overlap region of the filaments changes the movement of space charges in the plasma, thereby changing the angular distribution of THz radiation. As a result, this schematic induces some THz wave from the second filament to propagate along the path of the THz wave from the first filament. Thus, this schematic alters the direction of the THz radiation from the filamentation, which can be used in THz wave remote sensing.

Phage or foe: an insight into the impact of viral predation on microbial communities.

Science.gov (United States)

Fernández, Lucía; Rodríguez, Ana; García, Pilar

2018-05-01

Since their discovery, bacteriophages have been traditionally regarded as the natural enemies of bacteria. However, recent advances in molecular biology techniques, especially data from "omics" analyses, have revealed that the interplay between bacterial viruses and their hosts is far more intricate than initially thought. On the one hand, we have become more aware of the impact of viral predation on the composition and genetic makeup of microbial communities thanks to genomic and metagenomic approaches. Moreover, data obtained from transcriptomic, proteomic, and metabolomic studies have shown that responses to phage predation are complex and diverse, varying greatly depending on the bacterial host, phage, and multiplicity of infection. Interestingly, phage exposure may alter different phenotypes, including virulence and biofilm formation. The complexity of the interactions between microbes and their viral predators is also evidenced by the link between quorum-sensing signaling pathways and bacteriophage resistance. Overall, new data increasingly suggests that both temperate and virulent phages have a positive effect on the evolution and adaptation of microbial populations. From this perspective, further research is still necessary to fully understand the interactions between phage and host under conditions that allow co-existence of both populations, reflecting more accurately the dynamics in natural microbial communities.

Phages can constrain protist predation-driven attenuation of Pseudomonas aeruginosa virulence in multienemy communities

Science.gov (United States)

Friman, Ville-Petri; Buckling, Angus

2014-01-01

The coincidental theory of virulence predicts that bacterial pathogenicity could be a by-product of selection by natural enemies in environmental reservoirs. However, current results are ambiguous and the simultaneous impact of multiple ubiquitous enemies, protists and phages on virulence evolution has not been investigated previously. Here we tested experimentally how Tetrahymena thermophila protist predation and PNM phage parasitism (bacteria-specific virus) alone and together affect the evolution of Pseudomonas aeruginosa PAO1 virulence, measured in wax moth larvae. Protist predation selected for small colony types, both in the absence and presence of phage, which showed decreased edibility to protists, reduced growth in the absence of enemies and attenuated virulence. Although phage selection alone did not affect the bacterial phenotype, it weakened protist-driven antipredatory defence (biofilm formation), its associated pleiotropic growth cost and the correlated reduction in virulence. These results suggest that protist selection can be a strong coincidental driver of attenuated bacterial virulence, and that phages can constrain this effect owing to effects on population dynamics and conflicting selection pressures. Attempting to define causal links such as these might help us to predict the cold and hot spots of coincidental virulence evolution on the basis of microbial community composition of environmental reservoirs. PMID:24671085

Impact of reducing and oxidizing agents on the infectivity of Qβ phage and the overall structure of its capsid.

Science.gov (United States)

Loison, Pauline; Majou, Didier; Gelhaye, Eric; Boudaud, Nicolas; Gantzer, Christophe

2016-11-01

Qβ phages infect Escherichia coli in the human gut by recognizing F-pili as receptors. Infection therefore occurs under reducing conditions induced by physiological agents (e.g. glutathione) or the intestinal bacterial flora. After excretion in the environment, phage particles are exposed to oxidizing conditions and sometimes disinfection. If inactivation does not occur, the phage may infect new hosts in the human gut through the oral route. During such a life cycle, we demonstrated that, outside the human gut, cysteines of the major protein capsid of Qβ phage form disulfide bonds. Disinfection with NaClO does not allow overoxidation to occur. Such oxidation induces inactivation rather by irreversible damage to the minor proteins. In the presence of glutathione, most disulfide bonds are reduced, which slightly increases the capacity of the phage to infect E. coli in vitro Such reduction is reversible and barely alters infectivity of the phage. Reduction of all disulfide bonds by dithiothreitol leads to complete capsid destabilization. These data provide new insights into how the phages are impacted by oxidizing-reducing conditions outside their host cell and raises the possibility of the intervention of the redox during life cycle of the phage. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

A Genetic Approach to the Development of New Therapeutic Phages to Fight Pseudomonas Aeruginosa in Wound Infections

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Elena Pleteneva

2012-12-01

Full Text Available Pseudomonas aeruginosa is a frequent participant in wound infections. Emergence of multiple antibiotic resistant strains has created significant problems in the treatment of infected wounds. Phage therapy (PT has been proposed as a possible alternative approach. Infected wounds are the perfect place for PT applications, since the basic condition for PT is ensured; namely, the direct contact of bacteria and their viruses. Plenty of virulent (“lytic” and temperate (“lysogenic” bacteriophages are known in P. aeruginosa. However, the number of virulent phage species acceptable for PT and their mutability are limited. Besides, there are different deviations in the behavior of virulent (and temperate phages from their expected canonical models of development. We consider some examples of non-canonical phage-bacterium interactions and the possibility of their use in PT. In addition, some optimal approaches to the development of phage therapy will be discussed from the point of view of a biologist, considering the danger of phage-assisted horizontal gene transfer (HGT, and from the point of view of a surgeon who has accepted the Hippocrates Oath to cure patients by all possible means. It is also time now to discuss the possible approaches in international cooperation for the development of PT. We think it would be advantageous to make phage therapy a kind of personalized medicine.

Intermediate filament mechanics in vitro and in the cell: From coiled coils to filaments, fibers and networks

OpenAIRE

Köster, Sarah; Weitz, David; Goldman, Robert D.; Aebi, Ueli; Herrmann, Harald

2015-01-01

Intermediate filament proteins form filaments, fibers and networks both in the cytoplasm and the nucleus of metazoan cells. Their general structural building plan accommodates highly varying amino acid sequences to yield extended dimeric α-helical coiled coils of highly conserved design. These “rod” particles are the basic building blocks of intrinsically flexible, filamentous structures that are able to resist high mechanical stresses, i.e. bending and stretching to a considerable degree, bo...

A Genotypic Analysis of Five P. aeruginosa Strains after Biofilm Infection by Phages Targeting Different Cell Surface Receptors

Directory of Open Access Journals (Sweden)

Diana P. Pires

2017-06-01

Full Text Available Antibiotic resistance constitutes one of the most serious threats to the global public health and urgently requires new and effective solutions. Bacteriophages are bacterial viruses increasingly recognized as being good alternatives to traditional antibiotic therapies. In this study, the efficacy of phages, targeting different cell receptors, against Pseudomonas aeruginosa PAO1 biofilm and planktonic cell cultures was evaluated over the course of 48 h. Although significant reductions in the number of viable cells were achieved for both cases, the high level of adaptability of the bacteria in response to the selective pressure caused by phage treatment resulted in the emergence of phage-resistant variants. To further investigate the genetic makeup of phage-resistant variants isolated from biofilm infection experiments, some of these bacteria were selected for phenotypic and genotypic characterization. Whole genome sequencing was performed on five phage-resistant variants and all of them carried mutations affecting the galU gene as well as one of pil genes. The sequencing analysis further revealed that three of the P. aeruginosa PAO1 variants carry large deletions (>200 kbp in their genomes. Complementation of the galU mutants with wild-type galU in trans restored LPS expression on the bacterial cell surface of these bacterial strains and rendered the complemented strains to be sensitive to phages. This provides unequivocal evidence that inactivation of galU function was associated with resistance to the phages that uses LPS as primary receptors. Overall, this work demonstrates that P. aeruginosa biofilms can survive phage attack and develop phage-resistant variants exhibiting defective LPS production and loss of type IV pili that are well adapted to the biofilm mode of growth.

A novel Pseudomonas aeruginosa bacteriophage, Ab31, a chimera formed from temperate phage PAJU2 and P. putida lytic phage AF: characteristics and mechanism of bacterial resistance.

Directory of Open Access Journals (Sweden)

Libera Latino

Full Text Available A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31 is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10 with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung.

Self-assembly of designed supramolecular magnetic filaments of different shapes

Energy Technology Data Exchange (ETDEWEB)

Novak, E.V. [Ural Federal University, Lenin Av. 51, Ekaterinburg (Russian Federation); Rozhkov, D.A., E-mail: d.a.rozhkov@gmail.com [Ural Federal University, Lenin Av. 51, Ekaterinburg (Russian Federation); Sanchez, P.A. [University of Vienna, Sensengasse 8, Vienna (Austria); Kantorovich, S.S. [Ural Federal University, Lenin Av. 51, Ekaterinburg (Russian Federation); University of Vienna, Sensengasse 8, Vienna (Austria)

2017-06-01

In the present work we study via molecular dynamics simulations filaments of ring and linear shape. Filaments are made of magnetic nanoparticles, possessing a point dipole in their centres. Particles in filaments are crosslinked in a particular way, so that the deviation of the neighbouring dipoles from the head-to-tail orientation is penalised by the bond. We show how the conformation of a single chain and ring filament changes on cooling for different lengths. We also study filament pairs, by fixing filaments at a certain distance and analysing the impact of inter-filament interaction on the equilibrium configurations. Our study opens a perspective to investigate the dispersions of filaments, both theoretically and numerically, by using effective potentials. - Highlights: • Single filament study. • Magnetic particles crosslinked in chains and rings. • Magnetic filament interactions.

A comparison study of a solar active-region eruptive filament and a neighboring non-eruptive filament

Science.gov (United States)

Jiang, Chao-Wei; Wu, Shi-Tsan; Feng, Xue-Shang; Hu, Qiang

2016-01-01

Solar active region (AR) 11283 is a very magnetically complex region and it has produced many eruptions. However, there exists a non-eruptive filament in the plage region just next to an eruptive one in the AR, which gives us an opportunity to perform a comparison analysis of these two filaments. The coronal magnetic field extrapolated using our CESE-MHD-NLFFF code reveals that two magnetic flux ropes (MFRs) exist in the same extrapolation box supporting these two filaments, respectively. Analysis of the magnetic field shows that the eruptive MFR contains a bald-patch separatrix surface (BPSS) cospatial very well with a pre-eruptive EUV sigmoid, which is consistent with the BPSS model for coronal sigmoids. The magnetic dips of the non-eruptive MFRs match Hα observation of the non-eruptive filament strikingly well, which strongly supports the MFR-dip model for filaments. Compared with the non-eruptive MFR/filament (with a length of about 200 Mm), the eruptive MFR/filament is much smaller (with a length of about 20 Mm), but it contains most of the magnetic free energy in the extrapolation box and holds a much higher free energy density than the non-eruptive one. Both the MFRs are weakly twisted and cannot trigger kink instability. The AR eruptive MFR is unstable because its axis reaches above a critical height for torus instability, at which the overlying closed arcades can no longer confine the MFR stably. On the contrary, the quiescent MFR is very firmly held by its overlying field, as its axis apex is far below the torus-instability threshold height. Overall, this comparison investigation supports that an MFR can exist prior to eruption and the ideal MHD instability can trigger an MFR eruption.

A comparison study of a solar active-region eruptive filament and a neighboring non-eruptive filament

International Nuclear Information System (INIS)

Jiang, Chao-Wei; Feng, Xue-Shang; Wu, Shi-Tsan; Hu, Qiang

2016-01-01

Solar active region (AR) 11283 is a very magnetically complex region and it has produced many eruptions. However, there exists a non-eruptive filament in the plage region just next to an eruptive one in the AR, which gives us an opportunity to perform a comparison analysis of these two filaments. The coronal magnetic field extrapolated using our CESE–MHD–NLFFF code reveals that two magnetic flux ropes (MFRs) exist in the same extrapolation box supporting these two filaments, respectively. Analysis of the magnetic field shows that the eruptive MFR contains a bald-patch separatrix surface (BPSS) cospatial very well with a pre-eruptive EUV sigmoid, which is consistent with the BPSS model for coronal sigmoids. The magnetic dips of the non-eruptive MFRs match Hα observation of the non-eruptive filament strikingly well, which strongly supports the MFR-dip model for filaments. Compared with the non-eruptive MFR/filament (with a length of about 200 Mm), the eruptive MFR/filament is much smaller (with a length of about 20 Mm), but it contains most of the magnetic free energy in the extrapolation box and holds a much higher free energy density than the non-eruptive one. Both the MFRs are weakly twisted and cannot trigger kink instability. The AR eruptive MFR is unstable because its axis reaches above a critical height for torus instability, at which the overlying closed arcades can no longer confine the MFR stably. On the contrary, the quiescent MFR is very firmly held by its overlying field, as its axis apex is far below the torus-instability threshold height. Overall, this comparison investigation supports that an MFR can exist prior to eruption and the ideal MHD instability can trigger an MFR eruption. (paper)

Helical beating of an actuated elastic filament

International Nuclear Information System (INIS)

Coq, Nais; Roure, Olivia du; Fermigier, Marc; Bartolo, Denis

2009-01-01

We investigate the propulsive force resulting from the rotation of a flexible filament in the low Reynolds number regime. Using a simple linear model, we establish the nonlinear torque-force relations for two torque-driven actuation modes. When the rotation of the filament is induced by two perpendicular transverse oscillating torques, the propulsive force increases monotonically with the torque amplitude. Conversely, when a constant axial torque is applied, the torque-force characteristics displays an unstable branch, related to a discontinuous transition in the shape of the filament. We characterize this shape transition using two geometrical parameters, quantifying the wrapping around and the collapse on the axis of the filament. The proposed theoretical description correctly accounts for our experimental observations and reveals a strong dependence of the filament dynamics on the anchoring conditions.

W-reactivation of phage lambda in X-irradiated mutants of Escherichia coli K-12

Energy Technology Data Exchange (ETDEWEB)

Martignoni, K D; Haselbacher, I [Muenchen Univ. (Germany, F.R.). Strahlenbiologisches Inst.

1980-07-01

The survival of UV irradiated phage lambda was increased on X-irradiated E.coli K-12 host cells over that on unirradiated cells. The frequency of c mutants among the surviving phages was increased to a similar extent by the X-ray exposure of the host cells as by UV light. This W-reactivation of phage lambda occurred in uvrA, polA, and recB mutants besides the wild type at about equal X-ray doses, but at a reduced reactivation efficiency compared with the wild type. W-reactivation was undetectable in recA mutants. While maximal UV induced W-reactivation occured 30 min after irradiation, the maximal X-ray induced reactivation was found immediately after irradiation. Chloramphenicol (100 ..mu..g/ml) and nitrofurantoin (50 ..mu..g/ml) inhibited W-reactivation of phage lambda if added before irradiation of the host cells, indicating the necessity of protein synthesis for W-reactivation.

Assessment of the Effects of Various UV Sources on Inactivation and Photoproduct Induction in Phage T7 Dosimeter

NARCIS (Netherlands)

Fekete, A.; Vink, A.A.; Gaspar, S.; Berces, A.; Modos, K.; Ronto, Gy.; Roza, L.

1998-01-01

The correlation between the biologically effective dose (BED) of a phage T7 biological dosimeter and the induction of cyclobutane pyrimidine dimers (CPD) and (6-4) photoproducts ((6-4)PD) in the phage DNA was determined using seven various UV sources. The BED is the inactivation rate of phage T7

Fundamentals of Filament Interaction

Science.gov (United States)

2017-05-19

AFRL-AFOSR-VA-TR-2017-0110 FUNDAMENTALS OF FILAMENT INTERACTION Martin Richardson UNIVERSITY OF CENTRAL FLORIDA Final Report 06/02/2017 DISTRIBUTION...of Filament Interaction 5a. CONTRACT NUMBER 5b. GRANT NUMBER FA95501110001 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Martin Richardson 5d. PROJECT...NAME OF RESPONSIBLE PERSON Martin Richardson a. REPORT b. ABSTRACT c. THIS PAGE 19b. TELEPHONE NUMBER (Include area code) 407-823-6819 Standard Form

Filaments in simulations of molecular cloud formation

Energy Technology Data Exchange (ETDEWEB)

Gómez, Gilberto C.; Vázquez-Semadeni, Enrique [Centro de Radioastronomía y Astrofísica, Universidad Nacional Autónoma de México, Campus Morelia Apartado Postal 3-72, 58090 Morelia, Michoacán (Mexico)

2014-08-20

We report on the filaments that develop self-consistently in a new numerical simulation of cloud formation by colliding flows. As in previous studies, the forming cloud begins to undergo gravitational collapse because it rapidly acquires a mass much larger than the average Jeans mass. Thus, the collapse soon becomes nearly pressureless, proceeding along its shortest dimension first. This naturally produces filaments in the cloud and clumps within the filaments. The filaments are not in equilibrium at any time, but instead are long-lived flow features through which the gas flows from the cloud to the clumps. The filaments are long-lived because they accrete from their environment while simultaneously accreting onto the clumps within them; they are essentially the locus where the flow changes from accreting in two dimensions to accreting in one dimension. Moreover, the clumps also exhibit a hierarchical nature: the gas in a filament flows onto a main, central clump but other, smaller-scale clumps form along the infalling gas. Correspondingly, the velocity along the filament exhibits a hierarchy of jumps at the locations of the clumps. Two prominent filaments in the simulation have lengths ∼15 pc and masses ∼600 M {sub ☉} above density n ∼ 10{sup 3} cm{sup –3} (∼2 × 10{sup 3} M {sub ☉} at n > 50 cm{sup –3}). The density profile exhibits a central flattened core of size ∼0.3 pc and an envelope that decays as r {sup –2.5} in reasonable agreement with observations. Accretion onto the filament reaches a maximum linear density rate of ∼30 M {sub ☉} Myr{sup –1} pc{sup –1}.

Thermal and Chemical Evolution of Collapsing Filaments

Energy Technology Data Exchange (ETDEWEB)

Gray, William J. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Scannapieco, Evan [Arizona State Univ., Mesa, AZ (United States). School of Earth and Space Exploration

2013-01-15

Intergalactic filaments form the foundation of the cosmic web that connect galaxies together, and provide an important reservoir of gas for galaxy growth and accretion. Here we present very high resolution two-dimensional simulations of the thermal and chemical evolution of such filaments, making use of a 32 species chemistry network that tracks the evolution of key molecules formed from hydrogen, oxygen, and carbon. We study the evolution of filaments over a wide range of parameters including the initial density, initial temperature, strength of the dissociating UV background, and metallicity. In low-redshift, Z ≈ 0.1Z_⊙ filaments, the evolution is determined completely by the initial cooling time. If this is sufficiently short, the center of the filament always collapses to form dense, cold core containing a substantial fraction of molecules. In high-redshift, Z = 10^-3Z_⊙ filaments, the collapse proceeds much more slowly. This is due mostly to the lower initial temperatures, which leads to a much more modest increase in density before the atomic cooling limit is reached, making subsequent molecular cooling much less efficient. Finally, we study how the gravitational potential from a nearby dwarf galaxy affects the collapse of the filament and compare this to NGC 5253, a nearby starbusting dwarf galaxy thought to be fueled by the accretion of filament gas. In contrast to our fiducial case, a substantial density peak forms at the center of the potential. This peak evolves faster than the rest of the filament due to the increased rate at which chemical species form and cooling occur. We find that we achieve similar accretion rates as NGC 5253, but our two-dimensional simulations do not recover the formation of the giant molecular clouds that are seen in radio observations.

Assessment of the Microscreen phage-induction assay for screening hazardous wastes

Energy Technology Data Exchange (ETDEWEB)

Houk, V.S.; DeMarini, D.M.

1987-09-01

The Microscreen phage-induction assay, which quantitatively measures the induction of prophage lambda in Escherichia coli WP2s(lambda), was used to test 14 crude (unfractionated) hazardous industrial waste samples for genotoxic activity in the presence and absence of metabolic activation. Eleven of the 14 wastes induced prophage, and induction was observed at concentrations as low as 0.4 picograms per ml. Comparisons between the mutagenicity of these waste samples in Salmonella and their ability to induce prophage lambda indicate that the Microscreen phage-induction assay detected genotoxic activity in all but one of the wastes that were mutagenic in Salmonella. Moreover, the Microscreen assay detected as genotoxic 5 additional wastes that were not detected in the Salmonella assay. The applicability of the Microscreen phage-induction assay for screening hazardous wastes for genotoxic activity is discussed along with some of the problems associated with screening highly toxic wastes containing toxic volatile compounds.

Commensal E. coli Stx2 lysogens produce high levels of phages after spontaneous prophage induction

Directory of Open Access Journals (Sweden)

Hildegunn eIversen

2015-02-01

Full Text Available Enterohemorrhagic E. coli (EHEC is a food-borne pathogen that causes disease ranging from uncomplicated diarrhea to life-threatening hemolytic uremic syndrome (HUS and nervous system complications. Shiga toxin 2 (Stx2 is the major virulence factor of EHEC and is critical for development of HUS. The genes encoding Stx2 are carried by lambdoid bacteriophages and the toxin production is tightly linked to the production of phages during lytic cycle. It has previously been suggested that commensal E. coli could amplify the production of Stx2-phages and contribute to the severity of disease. In this study we examined the susceptibility of commensal E. coli strains to the Stx2-converting phage ϕ734, isolated from a highly virulent EHEC O103:H25 (NIPH-11060424. Among 38 commensal E. coli strains from healthy children below five years, 15 were lysogenized by the ϕ734 phage, whereas lytic infection was not observed. Three of the commensal E. coli ϕ734 lysogens were tested for stability, and appeared stable and retained the phage for at least 10 cultural passages. When induced to enter lytic cycle by H2O2 treatment, 8 out of 13 commensal lysogens produced more ϕ734 phages than NIPH-11060424. Strikingly, five of them even spontaneously (non-induced produced higher levels of phage than the H2O2 induced NIPH-11060424. An especially high frequency of HUS (60% was seen among children infected by NIPH-11060424 during the outbreak in 2006. Based on our findings, a high Stx2 production by commensal E. coli lysogens cannot be ruled out as a contributor to the high frequency of HUS during this outbreak.

Isolation and characterization of φkm18p, a novel lytic phage with therapeutic potential against extensively drug resistant Acinetobacter baumannii.

Directory of Open Access Journals (Sweden)

Gwan-Han Shen

Full Text Available AIMS: To isolate phages against extensively drug resistant Acinetobacter baumannii (XDRAB and characterize the highest lytic capability phage as a model to evaluate the potential on phage therapy. METHODS AND RESULTS: Eight phages were isolated from hospital sewage and showed narrow host spectrum. Phage φkm18p was able to effectively lyse the most XDRAB. It has a dsDNA genome of 45 kb in size and hexagonal head of about 59 nm in diameter and no tail. Bacterial population decreased quickly from 10(8 CFU ml(-1 to 10(3 CFU ml(-1 in 30 min by φkm18p. The 185 kDa lysis protein encoded by φkm18p genome was detected when the extracted protein did not boil before SDS-PAGE; it showed that the lysis protein is a complex rather than a monomer. Phage φkm18p improved human lung epithelial cells survival rates when they were incubated with A. baumannii. Combination of phages (φkm18p, φTZ1 and φ314 as a cocktail could lyse all genotype-varying XDRAB isolates. CONCLUSION: Infections with XDRAB are extremely difficult to treat and development of a phage cocktails therapy could be a therapeutic alternative in the future. Phage φkm18p is a good candidate for inclusion in phage cocktails.

Phage induction by UV and mitomycin C in Pseudomonas mori, the pathogen of bacterial blight of mulberry

International Nuclear Information System (INIS)

Sato, Mamoru

1979-01-01

Phage induction by ultraviolet radiation (UV) and mitomycin C (MMC) in some lysogenic strains of Pseudomonas mori, the pathogen of bacterial blight of mulberry, was examined. Among 5 strains tested, in the strains S 6804 and S 6805, phage was induced by both UV and MMC, and in the strain M 5, only by MMC. In the strains S 6807 and S 6808, it was not induced by both these inducers. The rate of phage production in the strain 6805 was highest when it was exposed to UV (15 W UV lamp, 40 cm) for 5 seconds, by which about 90% of the bacteria were killed, and decreased rapidly by further extending the exposure time. The bacteria suspended in 0.02 M magnesium solution were more sensitive in responding to UV than those suspended in nutrient broth, but after the UV treatment, nutrient broth was more favorable than magnesium solution for phage production. The MMC added to nutrient broth induced phage production at the concentration from 0.5 to 5 μg/ml. The strains induced by either UV or MMC their temperate phages after about 3 hours of latent period. The phage induction by UV was almost completely suppressed by 40 minute exposure to fluorescent light (a 15 W fluorescent lamp, 10 cm) or by 5 minute exposure to sunlight, given within 45 minutes after the UV treatment, i.e. within 1/4 of the latent period. Thus, the photoreversion of the UV effect on phage induction was observed in Ps. mori as well as in Ps. pyocyanea and E. coli. (Kaihara, S.)

Use of phages against antibiotic-resistant Staphylococcus aureus isolated from bovine mastitis.

Science.gov (United States)

Dias, R S; Eller, M R; Duarte, V S; Pereira, Â L; Silva, C C; Mantovani, H C; Oliveira, L L; Silva, E de A M; De Paula, S O

2013-08-01

Bovine mastitis is the primary disease of dairy cattle worldwide and it causes large economic losses. Among several microorganisms that are the causative agents of this disease, Staphylococcus aureus is the most prevalent. Although antibiotic therapy is still the most widely used procedure for the treatment of bovine mastitis, alternative means of treatment are necessary due to the presence of antibiotic residues in milk, which is a growing concern because of its interference with the production of milk derivatives and the selection of resistant bacterial strains. The use of bacteriophages as a tool for the control of pathogens is an alternative treatment to antibiotic therapy. In this work, to obtain phages with the potential for use in phage therapy as a treatment for mastitis, we isolated and identified the bacteria from the milk of mastitis-positive cows. A total of 19% of the animals from small and medium farms of the Zona da Mata Mineira, Brazil, was positive for bovine mastitis, and bacteria of the genus Staphylococcus were the most prevalent pathogens. The majority of the S. aureus isolates tested was resistant to penicillin and ampicillin. In parallel, we isolated 10 bacteriophages able to infect some of these S. aureus isolates. We determined that these phages contained DNA genomes of approximately 175 kb in length, and the protein profiles indicated the presence of 4 major proteins. Electron microscopy revealed that the phages are caudate and belong to the Myoviridae family. The isolates exhibited interesting features for their use in phage therapy such as a high lytic potential, a wide range of hosts, and thermostability, all of which favor their use in the field.

Phage display peptide libraries: deviations from randomness and correctives

Science.gov (United States)

Ryvkin, Arie; Ashkenazy, Haim; Weiss-Ottolenghi, Yael; Piller, Chen; Pupko, Tal; Gershoni, Jonathan M

2018-01-01

Abstract Peptide-expressing phage display libraries are widely used for the interrogation of antibodies. Affinity selected peptides are then analyzed to discover epitope mimetics, or are subjected to computational algorithms for epitope prediction. A critical assumption for these applications is the random representation of amino acids in the initial naïve peptide library. In a previous study, we implemented next generation sequencing to evaluate a naïve library and discovered severe deviations from randomness in UAG codon over-representation as well as in high G phosphoramidite abundance causing amino acid distribution biases. In this study, we demonstrate that the UAG over-representation can be attributed to the burden imposed on the phage upon the assembly of the recombinant Protein 8 subunits. This was corrected by constructing the libraries using supE44-containing bacteria which suppress the UAG driven abortive termination. We also demonstrate that the overabundance of G stems from variant synthesis-efficiency and can be corrected using compensating oligonucleotide-mixtures calibrated by mass spectroscopy. Construction of libraries implementing these correctives results in markedly improved libraries that display random distribution of amino acids, thus ensuring that enriched peptides obtained in biopanning represent a genuine selection event, a fundamental assumption for phage display applications. PMID:29420788

Ultraviolet treatment on high performance filaments

International Nuclear Information System (INIS)

Gu Huang

2005-01-01

Quartz, Kevlar, carbon, and glass filaments were irradiated by ultraviolet ray with various periods. Tensile strength of the treated fibres was tested and analyzed, and the outward appearance of the treated filaments was shown

Observations of the Growth of an Active Region Filament

Science.gov (United States)

Yang, Bo

2017-04-01

We present observations of the growth of an active region filament caused by magnetic interactions among the filament and its adjacent superpenumbral filament (SF) and dark thread-like structures (T). Multistep reconnections are identified during the whole growing process. Magnetic flux convergence and cancellation occurring at the positive footpoint region of the filament is the first step reconnection, which resulted in the filament bifurcating into two sets of intertwined threads. One set anchored in situ, while the other set moved toward and interacted with the SF and part of T. This indicates the second step reconnection, which gave rise to the disappearance of the SF and the formation of a long thread-like structure that connects the far ends of the filament and T. The long thread-like structure further interacted with the T and then separated into two parts, representing the third step reconnection. Finally, another similar long thread-like structure, which intertwined with the fixed filament threads, appeared. Hαobservations show that this twisted structure is a longer sinistral filament. Based on the observed photospheric vector magnetograms, we performed a non-linear force-free field extrapolation to reconstruct the magnetic fields above the photosphere and found that the coronal magnetic field lines associated with the filament consists of two twisted flux ropes winding around each other. These results suggest that magnetic interactions among filaments and their adjacent SFs and T could lead to the growth of the filaments, and the filament is probably supported in a flux rope.

Identification of keratinocyte specific markers using phage display and mass spectrometry

DEFF Research Database (Denmark)

Jensen, K.B.; Jensen, O.N.; Ravn, P.

2003-01-01

and mass spectrometry that allows identification of cell type-specific protein markers. The most important features of the method are (i) reduction of experimental noise originating from background binding of phage particles and (ii) isolation of affinity binders after a single round of selection, which...... antigens were subsequently identified by mass spectrometry as laminin-5, plectin, and fibronectin. The combination of phage display technology with mass spectrometry methods for protein identification is a general and promising approach for proteomic analysis of cell surface complexity....

Removal of phages and viral pathogens in a full-scale MBR: Implications for wastewater reuse and potable water.

Science.gov (United States)

Purnell, Sarah; Ebdon, James; Buck, Austen; Tupper, Martyn; Taylor, Huw

2016-09-01

The aim of this study was to demonstrate how seasonal variability in the removal efficacy of enteric viral pathogens from an MBR-based water recycling system might affect risks to human health if the treated product were to be used for the augmentation of potable water supplies. Samples were taken over a twelve month period (March 2014-February 2015), from nine locations throughout a water recycling plant situated in East London and tested for faecal indicator bacteria (thermotolerant coliforms, intestinal enterococci n = 108), phages (somatic coliphage, F-specific RNA phage and Bacteroides phage (GB-124) n = 108), pathogenic viruses (adenovirus, hepatitis A, norovirus GI/GII n = 48) and a range of physico-chemical parameters (suspended solids, DO, BOD, COD). Thermotolerant coliforms and intestinal enterococci were removed effectively by the water recycling plant throughout the study period. Significant mean log reductions of 3.9-5.6 were also observed for all three phage groups monitored. Concentrations of bacteria and phages did not vary significantly according to season (P < 0.05; Kruskal-Wallis), though recorded levels of norovirus (GI) were significantly higher during autumn/winter months (P = 0.027; Kruskal-Wallis). Log reduction values for norovirus and adenovirus following MBR treatment were 2.3 and 4.4, respectively. However, both adenovirus and norovirus were detected at low levels (2000 and 3240 gene copies/L, respectively) post chlorination in single samples. Whilst phage concentrations did correlate with viral pathogens, the results of this study suggest that phages may not be suitable surrogates, as viral pathogen concentrations varied to a greater degree seasonally than did the phage indicators and were detected on a number of occasions on which phages were not detected (false negative sample results). Copyright © 2016 Elsevier Ltd. All rights reserved.

Colored fused filament fabrication

OpenAIRE

Song, Haichuan; Lefebvre, Sylvain

2017-01-01

Filament fused fabrication is the method of choice for printing 3D models at low cost, and is the de-facto standard for hobbyists, makers and schools. Unfortunately, filament printers cannot truly reproduce colored objects. The best current techniques rely on a form of dithering exploiting occlusion, that was only demonstrated for shades of two base colors and that behaves differently depending on surface slope. We explore a novel approach for 3D printing colored objects, capable of creating ...

Covalent and non-covalent chemical engineering of actin for biotechnological applications.

Science.gov (United States)

Kumar, Saroj; Mansson, Alf

2017-11-15

The cytoskeletal filaments are self-assembled protein polymers with 8-25nm diameters and up to several tens of micrometres length. They have a range of pivotal roles in eukaryotic cells, including transportation of intracellular cargoes (primarily microtubules with dynein and kinesin motors) and cell motility (primarily actin and myosin) where muscle contraction is one example. For two decades, the cytoskeletal filaments and their associated motor systems have been explored for nanotechnological applications including miniaturized sensor systems and lab-on-a-chip devices. Several developments have also revolved around possible exploitation of the filaments alone without their motor partners. Efforts to use the cytoskeletal filaments for applications often require chemical or genetic engineering of the filaments such as specific conjugation with fluorophores, antibodies, oligonucleotides or various macromolecular complexes e.g. nanoparticles. Similar conjugation methods are also instrumental for a range of fundamental biophysical studies. Here we review methods for non-covalent and covalent chemical modifications of actin filaments with focus on critical advantages and challenges of different methods as well as critical steps in the conjugation procedures. We also review potential uses of the engineered actin filaments in nanotechnological applications and in some key fundamental studies of actin and myosin function. Finally, we consider possible future lines of investigation that may be addressed by applying chemical conjugation of actin in new ways. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Bad Phages in Good Bacteria: Role of the Mysterious orf63 of λ and Shiga Toxin-Converting Φ24B Bacteriophages

Directory of Open Access Journals (Sweden)

Aleksandra Dydecka

2017-08-01

Full Text Available Lambdoid bacteriophages form a group of viruses that shares a common schema of genome organization and lifecycle. Some of them can play crucial roles in creating the pathogenic profiles of Escherichia coli strains. For example, Shiga toxin-producing E. coli (STEC acquired stx genes, encoding Shiga toxins, via lambdoid prophages (Stx phages. The results obtained so far present the evidence for the relation between the exo-xis region of the phage genome and lambdoid phage development, however molecular mechanisms of activities of the exo-xis genes' products are still unknown. In view of this, we decided to determine the influence of the uncharacterized open reading frame orf63 of the exo-xis region on lambdoid phages development using recombinant prophages, λ and Stx phage Φ24B. We have demonstrated that orf63 codes for a folded protein, thus, it is a functional gene. NMR spectroscopy and analytical gel filtration were used to extend this observation further. From backbone chemical shifts, Orf63 is oligomeric in solution, likely a trimer and consistent with its small size (63 aa., is comprised of two helices, likely intertwined to form the oligomer. We observed that the deletion of phage orf63 does not impair the intracellular lambdoid phage lytic development, however delays the time and decreases the efficiency of prophage induction and in consequence results in increased survival of E. coli during phage lytic development. Additionally, the deletion of phage orf63 negatively influences expression of the major phage genes and open reading frames from the exo-xis region during prophage induction with hydrogen peroxide. We conclude, that lambdoid phage orf63 may have specific functions in the regulation of lambdoid phages development, especially at the stage of the lysis vs. lysogenization decision. Besides, orf63 probably participates in the regulation of the level of expression of essential phage genes and open reading frames from the exo

Campylobacter jejuni motility is required for infection of the flagellotropic bacteriophage F341

DEFF Research Database (Denmark)

Baldvinsson, Signe Berg; Sørensen, Martine Camilla Holst; Vegge, Christina Skovgaard

2014-01-01

. In contrast, phage F341 does not infect C. jejuni NCTC11168 mutants that either lack the flagellar filaments (ΔflaAB) or that have paralyzed, i.e., nonrotating, flagella (ΔmotA and ΔflgP). Complementing flgP confirmed that phage F341 requires rotating flagella for successful infection. Furthermore, adsorption...

Biological control of Aeromonas salmonicida subsp. salmonicida infection in rainbow trout (Oncorhynchus mykiss) using Aeromonas phage PAS-1.

Science.gov (United States)

Kim, J H; Choresca, C H; Shin, S P; Han, J E; Jun, J W; Park, S C

2015-02-01

The potential control efficacy of Aeromonas phage PAS-1 was evaluated against Aeromonas salmonicida subsp. salmonicida infection in rainbow trout (Oncorhynchus mykiss) model in this study. The phage was co-cultured with the virulent A. salmonicida subsp. salmonicida strain AS05 that possesses the type III secretion system (TTSS) ascV gene, and efficient bacteriolytic activity was observed against the bacteria. The administration of PAS-1 in rainbow trout demonstrated that the phage was cleared from the fish within 200 h post-administration, and a temporal neutralizing activity against the phage was detected in the sera of phage-administrated fish. The administration of PAS-1 (multiplicity of infection: 10 000) in A. salmonicida subsp. salmonicida infected rainbow trout model showed notable protective effects, with increased survival rates and mean times to death. These results demonstrated that Aeromonas phage PAS-1 could be considered as an alternative biological control agent against A. salmonicida subsp. salmonicida infections in rainbow trout culture. © 2013 Blackwell Verlag GmbH.

Positrusion Filament Recycling System, Phase I

Data.gov (United States)

National Aeronautics and Space Administration — TUI proposes a novel process to produce 3d printer feedstock filament out of scrap ABS on the ISS. Currently the plastic filament materials that most 3d printers use...

Probing ADAMTS13 Substrate Specificity using Phage Display

Science.gov (United States)

Desch, Karl C.; Kretz, Colin; Yee, Andrew; Gildersleeve, Robert; Metzger, Kristin; Agrawal, Nidhi; Cheng, Jane; Ginsburg, David

2015-01-01

Von Willebrand factor (VWF) is a large, multimeric protein that regulates hemostasis by tethering platelets to the subendothelial matrix at sites of vascular damage. The procoagulant activity of plasma VWF correlates with the length of VWF multimers, which is proteolytically controlled by the metalloprotease ADAMTS13. To probe ADAMTS13 substrate specificity, we created phage display libraries containing randomly mutated residues of a minimal ADAMTS13 substrate fragment of VWF, termed VWF73. The libraries were screened for phage particles displaying VWF73 mutant peptides that were resistant to proteolysis by ADAMTS13. These peptides exhibited the greatest mutation frequency near the ADAMTS13 scissile residues. Kinetic assays using mutant and wild-type substrates demonstrated excellent agreement between rates of cleavage for mutant phage particles and the corresponding mutant peptides. Cleavage resistance of selected mutations was tested in vivo using hydrodynamic injection of corresponding full-length expression plasmids into VWF-deficient mice. These studies confirmed the resistance to cleavage resulting from select amino acid substitutions and uncovered evidence of alternate cleavage sites and recognition by other proteases in the circulation of ADAMTS13 deficient mice. Taken together, these studies demonstrate the key role of specific amino acids residues including P3-P2’ and P11’, for substrate specificity and emphasize the importance in flowing blood of other ADAMTS13–VWF exosite interactions outside of VWF73. PMID:25849793

Hydrodynamic interaction induced spontaneous rotation of coupled active filaments.

Science.gov (United States)

Jiang, Huijun; Hou, Zhonghuai

2014-12-14

We investigate the coupled dynamics of active filaments with long range hydrodynamic interactions (HI). Remarkably, we find that filaments can rotate spontaneously under the same conditions in which a single filament alone can only move in translation. Detailed analysis reveals that the emergence of coupled rotation originates from an asymmetric flow field associated with HI which breaks the symmetry of translational motion when filaments approach. The breaking is then further stabilized by HI to form self-sustained coupled rotation. Intensive simulations show that coupled rotation forms easily when one filament tends to collide with the front-half of the other. For head-to-tail approaching, we observe another interesting HI-induced coupled motion, where filaments move together in the form of one following the other. Moreover, the radius of coupled rotation increases exponentially as the rigidity of the filament increases, which suggests that HI are also important for the alignment of rigid-rod-like filaments which has been assumed to be solely a consequence of direct collisions.

Star-forming Filament Models

International Nuclear Information System (INIS)

Myers, Philip C.

2017-01-01

New models of star-forming filamentary clouds are presented in order to quantify their properties and to predict their evolution. These 2D axisymmetric models describe filaments that have no core, one low-mass core, and one cluster-forming core. They are based on Plummer-like cylinders and spheroids that are bounded by a constant-density surface of finite extent. In contrast to 1D Plummer-like models, they have specific values of length and mass, they approximate observed column density maps, and their distributions of column density ( N -pdfs) are pole-free. Each model can estimate the star-forming potential of a core-filament system by identifying the zone of gas dense enough to form low-mass stars and by counting the number of enclosed thermal Jeans masses. This analysis suggests that the Musca central filament may be near the start of its star-forming life, with enough dense gas to make its first ∼3 protostars, while the Coronet filament is near the midpoint of its star formation, with enough dense gas to add ∼8 protostars to its ∼20 known stars. In contrast, L43 appears to be near the end of its star-forming life, since it lacks enough dense gas to add any new protostars to the two young stellar objectsalready known.

Star-forming Filament Models

Energy Technology Data Exchange (ETDEWEB)

Myers, Philip C., E-mail: pmyers@cfa.harvard.edu [Harvard-Smithsonian Center for Astrophysics, 60 Garden Street, Cambridge, MA 02138 (United States)

2017-03-20

New models of star-forming filamentary clouds are presented in order to quantify their properties and to predict their evolution. These 2D axisymmetric models describe filaments that have no core, one low-mass core, and one cluster-forming core. They are based on Plummer-like cylinders and spheroids that are bounded by a constant-density surface of finite extent. In contrast to 1D Plummer-like models, they have specific values of length and mass, they approximate observed column density maps, and their distributions of column density ( N -pdfs) are pole-free. Each model can estimate the star-forming potential of a core-filament system by identifying the zone of gas dense enough to form low-mass stars and by counting the number of enclosed thermal Jeans masses. This analysis suggests that the Musca central filament may be near the start of its star-forming life, with enough dense gas to make its first ∼3 protostars, while the Coronet filament is near the midpoint of its star formation, with enough dense gas to add ∼8 protostars to its ∼20 known stars. In contrast, L43 appears to be near the end of its star-forming life, since it lacks enough dense gas to add any new protostars to the two young stellar objectsalready known.

Genome of a SAR116 bacteriophage shows the prevalence of this phage type in the oceans.

Science.gov (United States)

Kang, Ilnam; Oh, Hyun-Myung; Kang, Dongmin; Cho, Jang-Cheon

2013-07-23

The abundance, genetic diversity, and crucial ecological and evolutionary roles of marine phages have prompted a large number of metagenomic studies. However, obtaining a thorough understanding of marine phages has been hampered by the low number of phage isolates infecting major bacterial groups other than cyanophages and pelagiphages. Therefore, there is an urgent requirement for the isolation of phages that infect abundant marine bacterial groups. In this study, we isolated and characterized HMO-2011, a phage infecting a bacterium of the SAR116 clade, one of the most abundant marine bacterial lineages. HMO-2011, which infects "Candidatus Puniceispirillum marinum" strain IMCC1322, has an ~55-kb dsDNA genome that harbors many genes with novel features rarely found in cultured organisms, including genes encoding a DNA polymerase with a partial DnaJ central domain and an atypical methanesulfonate monooxygenase. Furthermore, homologs of nearly all HMO-2011 genes were predominantly found in marine metagenomes rather than cultured organisms, suggesting the novelty of HMO-2011 and the prevalence of this phage type in the oceans. A significant number of the viral metagenome sequences obtained from the ocean surface were best assigned to the HMO-2011 genome. The number of reads assigned to HMO-2011 accounted for 10.3%-25.3% of the total reads assigned to viruses in seven viromes from the Pacific and Indian Oceans, making the HMO-2011 genome the most or second-most frequently assigned viral genome. Given its ability to infect the abundant SAR116 clade and its widespread distribution, Puniceispirillum phage HMO-2011 could be an important resource for marine virus research.

Isolation and characterization of ZZ1, a novel lytic phage that infects Acinetobacter baumannii clinical isolates

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Jin Jing

2012-07-01

Full Text Available Abstract Background Acinetobacter baumannii, a significant nosocomial pathogen, has evolved resistance to almost all conventional antimicrobial drugs. Bacteriophage therapy is a potential alternative treatment for multidrug-resistant bacterial infections. In this study, one lytic bacteriophage, ZZ1, which infects A. baumannii and has a broad host range, was selected for characterization. Results Phage ZZ1 and 3 of its natural hosts, A. baumanni clinical isolates AB09V, AB0902, and AB0901, are described in this study. The 3 strains have different sensitivities to ZZ1, but they have the same sensitivity to antibiotics. They are resistant to almost all of the antibiotics tested, except for polymyxin. Several aspects of the life cycle of ZZ1 were investigated using the sensitive strain AB09V under optimal growth conditions. ZZ1 is highly infectious with a short latent period (9 min and a large burst size (200 PFU/cell. It exhibited the most powerful antibacterial activity at temperatures ranging from 35°C to 39°C. Moreover, when ZZ1 alone was incubated at different pHs and different temperatures, the phage was stable over a wide pH range (4 to 9 and at extreme temperatures (between 50°C and 60°C. ZZ1 possesses a 100-nm icosahedral head containing double-stranded DNA with a total length of 166,682 bp and a 120-nm long contractile tail. Morphologically, it could be classified as a member of the Myoviridae family and the Caudovirales order. Bioinformatic analysis of the phage whole genome sequence further suggested that ZZ1 was more likely to be a new member of the Myoviridae phages. Most of the predicted ORFs of the phage were similar to the predicted ORFs from other Acinetobacter phages. Conclusion The phage ZZ1 has a relatively broad lytic spectrum, high pH stability, strong heat resistance, and efficient antibacterial potential at body temperature. These characteristics greatly increase the utility of this phage as an antibacterial agent

Isolation and characterization of ZZ1, a novel lytic phage that infects Acinetobacter baumannii clinical isolates.

Science.gov (United States)

Jin, Jing; Li, Zhen-Jiang; Wang, Shu-Wei; Wang, Shan-Mei; Huang, De-Hai; Li, Ya-Hui; Ma, Yun-Yun; Wang, Jin; Liu, Fang; Chen, Xiang-Dong; Li, Guang-Xing; Wang, Xiao-Ting; Wang, Zhong-Quan; Zhao, Guo-Qiang

2012-07-28

Acinetobacter baumannii, a significant nosocomial pathogen, has evolved resistance to almost all conventional antimicrobial drugs. Bacteriophage therapy is a potential alternative treatment for multidrug-resistant bacterial infections. In this study, one lytic bacteriophage, ZZ1, which infects A. baumannii and has a broad host range, was selected for characterization. Phage ZZ1 and 3 of its natural hosts, A. baumanni clinical isolates AB09V, AB0902, and AB0901, are described in this study. The 3 strains have different sensitivities to ZZ1, but they have the same sensitivity to antibiotics. They are resistant to almost all of the antibiotics tested, except for polymyxin. Several aspects of the life cycle of ZZ1 were investigated using the sensitive strain AB09V under optimal growth conditions. ZZ1 is highly infectious with a short latent period (9 min) and a large burst size (200 PFU/cell). It exhibited the most powerful antibacterial activity at temperatures ranging from 35°C to 39°C. Moreover, when ZZ1 alone was incubated at different pHs and different temperatures, the phage was stable over a wide pH range (4 to 9) and at extreme temperatures (between 50°C and 60°C). ZZ1 possesses a 100-nm icosahedral head containing double-stranded DNA with a total length of 166,682 bp and a 120-nm long contractile tail. Morphologically, it could be classified as a member of the Myoviridae family and the Caudovirales order. Bioinformatic analysis of the phage whole genome sequence further suggested that ZZ1 was more likely to be a new member of the Myoviridae phages. Most of the predicted ORFs of the phage were similar to the predicted ORFs from other Acinetobacter phages. The phage ZZ1 has a relatively broad lytic spectrum, high pH stability, strong heat resistance, and efficient antibacterial potential at body temperature. These characteristics greatly increase the utility of this phage as an antibacterial agent; thus, it should be further investigated.