1,721,377 research outputs found
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Functionele ontrafeling van antibacteriële faageiwitten tegen Pseudomonas aeruginosa
No full textPseudomonas aeruginosa causes life-threatening infections. Considering the current poor rate of release of novel antibiotics, let alone entirely novel classes of antibiotics, it is a worrying indication that we may soon run out of treatment options. Therefore, the development of innovative antibiotics targeting (not yet exploited) essential bacterial pathways will be crucial inthe near future. Strictly lytic bacteriophages, bacterias natural enemies, rely completely on the bacterial metabolism for their propagation. Over a billion years of co-evolutionary struggle phages have evolved an incredible number of highly diverse proteins that either inhibit or adapt bacterialmetabolic processes to their own benefit. Many of them lead to cell-cycle arrest or even host lethality. As such, a novel source of Gram-negative antibacterials might originate from mining the thousands of available sequenced phage genomes. 158 early phage proteins encoded by nine different P. aeruginosa phages were selected as starting point of this work. We hypothesized that phage proteins, which are growth-inhibitory to their host when individually expressed, show the most promise in tackling crucial metabolic pathways. Consequently, the 158 selected proteins were first screened for their effect on P. aeruginosa growth. In total, nineteen unknown antibacterial phage proteins could be identified.To explore their possible mode of action and the molecular background of their toxicity, a systematic yeast two-hybrid (Y2H) against a random genomic fragment library of P. aeruginosa PAO1 was applied to identify their target(s) in Pseudomonas. This showed that bacteriophages influence the host metabolism using a variety of modes.A nice example is LUZ24 gp4. For this phage protein, one potential interaction partner in P. aeruginosa was identified, the PA4315-encoded transcriptional regulator MvaT, which was confirmed in vitro using coprecipitation assays. MvaT is a histone-like nucleoid structuring protein, which exerts a crucial role in compaction of the bacterial chromosome by the formation of oligomers. Moreover, the polymerization of the protein across AT-rich DNA strands, permits gene silencing of foreign DNA, thereby limiting any potentially adverse effects of such DNA. Recombinant MvaT-His and LUZ24 gp4-Strep were tested in gel shift assays, which proved the inhibitory effect of LUZ24 gp4 on MvaT DNA-binding activity. We therefore termed this gene product as Mip, the MvaT-inhibiting protein. A hypothesis on the biological role of Mip, one of the first proteins produced right after infection, can be made: Mip indeed prevents the AT-rich LUZ24 DNA from being physically blocked by MvaT oligomers right after its injection in the host cell. This strategy gives the phage a clear advantage since a physical blockage of its DNA rightafter injection, will not complete its infection cycle. Inhibition of MvaT by a phage-encoded protein will keep the phage DNA MvaT-free, thereby allowing phage transcription and thus completion of the phage infection cycle. Although microbial resistance is probably an unavoidable consequence of antibiotic therapy, a bacteriophage-based platform has a great potential with respect to identifying novel mechanisms and targets to treat bacterial infections. In fact, known phage-host interactions illustrate the potential for phage systems to be used for the identification of points in host metabolism that may be susceptible to small-molecule inhibitors. The most efficient and vulnerable targets are selected and validated through billions of years of co-evolution between phages and their hosts. As there is no dearth of bacteriophages in nature, the quest for lethal phage proteins as well as their cognate bacterial targets should be continued in order to expedite the research on antibacterial drug discovery.status: Publishe
Ontwikkeling en evaluatie van geoptimaliseerde bacteriofaagendolysines ter inactivatie van gramnegatieve bacteriën.
No full textBacteriophages, viruses infecting bacteria, disrupt the bacterial cell wall at the end of their replication cycle to release newly produced virions. The major constituent of the bacterial cell wall is the peptidoglycan. To degrade this rigid layer, bacteriophages encode peptidoglycan hydrolases, called endolysins, that hydrolyze specific bonds in the peptidoglycan. This dissertation specifically focuses on endolysins, isolated from phages infecting Gram-negative bacterial species, including Pseudomonas aeruginosa, Salmonella Typhimurium, Escherichia coli, Klebsiella pneumoniae and Citrobacter rodentii. Most of these bacteria are opportunistic pathogens that are of increasing concern in hospitals due to their high intrinsic and acquired antibiotic resistance. In a first part of this study, we extend the pool of available endolysins from Gram-negative origin and analyze their potential and applicability as alternative antibacterial agents for antibiotics to combat these Gram-negative pathogens. The Gram-negative outer membrane prevents exogenously applied endolysins from reaching the peptidoglycan layer and protects bacteria against their lytic activity. We therefore evaluate in the second part of this dissertation an approach that allows the endolysin to efficiently destabilize the outer membrane and subsequently reach the peptidoglycan. This approach consists of the fusion of a set of outer membrane-permeabilizing antimicrobial peptides to the endolysin to allow for an autonomous interaction with the outer membrane. To sketch the background, this dissertation starts with an overview of the literature concerning bacteriophage endolysins (history and structural diversity), antimicrobial peptides (types and mode of action) and outer membrane diversity present among Gram-negative bacteria.From an in silico analysis of fully sequenced phage genomes, a selection of fifteen interesting candidate endolysins is made (Chapter 4). Six single-domain (Chapter 5) and three modular (Chapter 6) endolysins with the highest maximal muralytic activity under physiological conditions, are selected for extensive characterization on biochemical (pH-dependency, enzymatic activity, activity upon heating) and antibacterial level. In this way, we aim to prove their lytic role and to reveal enzyme-specific characteristics interesting from an application perspective. In silico, the single-domain endolysins consist of a catalytic domain, whereas the modular ones feature an N-terminal peptidoglycan binding domain and a C-terminal catalytic domain, hitherto a unique property present in a few endolysins from Gram-negative origin. In addition, the predicted peptidoglycan binding domains are experimentally verified.The modular endolysins in this study are shown to be enzymatically more active than the single-domain endolysins, an observation that was translated into their in vitro antibacterial activity. Of all tested endolysins, the modular endolysin from Pseudomonas fluorescens phage OBP, OBPgp279, shows the highest muralytic and antibacterial activity, followed by PVP-SE1gp146, the endolysin from Salmonella Enteritidis phage PVP-SE1. The peptidoglycan binding domain present in modular endolysins accounts for their strong lytic action since the contribution of this domain (38 to 56 %) to the total enzymatic activity is considerable. In addition, the enzymatic activity is consistent for the different Gram-negative bacterial species due to their conserved peptidoglycan (A1gamma chemotype). This characterization also revealed various interesting biochemical properties. OBPgp279 shows intrinsic antibacterial activity on P. aeruginosa PAO1 (± 1 log unit), probably by destabilizing the Pseudomonas outer membrane. PVP-SE1gp146 remains active up to temperatures of 90°C with 60 % residual enzymatic activity after 40 minutes. This last property makes the enzyme a potential candidate as antibacterial component in hurdle technology for food preservation. At the start of the second part, OBPgp279 and PVP-SE1gp146, the two most promising endolysins, are selected to evaluate the proposed fusion approach for passage of the outer membrane (Chapter 7). The N-terminal fusion of a polycationic PK peptide (KRKKRKKRK) composed of lysine and arginine residues, turns out to be the most effective fusion to improve the antibacterial activity of both endolysins. The highest activity is reached for P. aeruginosa with maximal 2.61 log units. Addition of minor EDTA concentrations enhances activity and extends the activity range with E. coli (maximal 1.70 log units) and S. Typhimurium (maximal 0.91 log units). This fused PK peptide is believed to compete with the Achilles heel of the outer membrane: the stabilizing divalent cations. A double N-terminal fusion of this promising PK peptide with other antimicrobial peptides only increases the antibacterial activity of OBPgp279 against E. coli to maximal 2.22 log units (for PP-PK double fusion), but is detrimental for the activity against other Gram-negative species. Analysis for the impact of the N-terminal PK fusion on endolysin characteristics reveals a protein-dependent inhibition of the enzymatic activity (with 52 to 94 %), a reduced heat resistance and a switch in pH-dependency to slightly more alkaline values (Chapter 8). Due to a more hydrophobic outer membrane, the antibacterial efficacy of the PK fusion is limited for Enterobacteriaceae. Additionally, the PK fusion also confers biofilm-degrading activity to PVP-SE1gp146. Extension of the linker length between the PK peptide and endolysin partly reconstitutes the reduced muralytic activity due to the PK peptide fusion, leading to an improved antibacterial activity against Pseudomonads and Enterobacteriaceae. Switching the PK peptide to the C-terminal end does not improve activity. These data nicely illustrate that endolysins can be turned into effective anti-Gram-negative compounds by an N-terminalfusion approach and subsequent optimization of the linker length.In the last part, we evaluate the PK-fused endolysin approach on an in vitro human keratinocyte monolayer and an in vivo Caenorhabditis elegans model. PK-PVP-SE1gp146 is able to protect the keratinocyte monolayer from a P. aeruginosa PA14 infection (Chapter 9). In addition, PK-PVP-SE1gp146 improves the survival of PA14-infected C. elegans with 60 % after five days of treatment (Chapter 10). These results prove the in vitro and in vivo applicability of the PK-fused endolysin approach against P. aeruginosa, offering promising perspectives towards prophylactic and therapeutic applications in human health and veterinary and towards microbial decontamination purposes in the food industry.status: Publishe
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
Karakterisering van Pseudomonas-infecterende bacteriofagen: de zoektocht naar virusgeassocieerde biofilmdegraderende enzymen
No full textThe biofilm mode of growth represents an important bacterial survival strategy, providing the enclosed bacterial cells with an increased resistance to antimicrobial agents and to the host immune system. Biofilm formation has been implicated in more than 60% of all bacterial infections where they form a source of recurrent infections. Moreover, it is one of the many bacterial resistance mechanisms to bacterial viruses, (bacterio)phages, their natural predators. In their co-evolutionary arms race, particular phages have developed the ability to tunnel through these biofilms. These phage posses an enzymatic activity in their virion-associated tail spikes or fibers which degrades the bacterial exopolysaccharides (EPS), one of the main constituting components of the biofilm matrix. These phages form a promising tool with regard to the renewed interest for phage therapy.The Pseudomonas infecting phages and their associated tail spikes or fibers form the main objective of this dissertation. Based on the formation of the typical plaque morphology with increasing opaque-looking halo zones, phages with potential virion-associated EPS depolymerase activity were selected and subjected to an in-depth characterization of their biofilm degradative properties. The 283,757 bp dsDNA genome of Pseudomonas fluorescens phage OBP possesses strong sequence similarity to the genome of Pseudomonas phage EL. Comparison of the genomic organization of the φKZ-related phages assembled in syntenic genomic blocks interspersed with hyperplastic regions, supports the proposed division in the EL-like viruses and the phiKZ-like viruses within a larger subfamily. Identification of putative early transcriptional promoters scattered throughout the hyperplastic regions, explains several features of the φKZ-related genome organization (existence of genomic islands) and evolution (multi-inversion in hyperplastic regions). Using Hidden Markov modeling typical conserved core genes encoding the portal protein, the injection needle, and two polypeptides with similarity to the 3-5 exonuclease domain and the polymerase domain of the T4 DNA polymerase, respectively, were identified. Two putative OBP paralog families possibly coding for the abundant proteinaceous fibers which are attached to the OBP tail structure, were annotated. While the N-terminal domains of the peptidoglycan degrading proteins and of one of these tail fiber modules are conserved, the observation of C-terminal catalytic domains typical for the different genera supports the further subdivision of the φKZ-related phages in the two separate genera. The Pseudomonas putida phages AF and φ15 were selected in collaboration with Prof. V.N. Krylov (Laboratory for Genetics of Bacteriophages, Russian Academy of Medical Sciences, Russia) as they form expanding halo zones around the zone of infection. Although φ15 and AF are related to different genera - the T7-like viruses and the epsilon15-like viruses andBPP-1-like viruses, respectively - of the Podoviridae family, both phages possess a closely related EPS-degrading tail spike protein which assembles in a SDS-resistant trimer. This tail spike protein appears to be the sole viral structural component which determines host specificity, since both phages share an identical host spectrum on a library of 53 P. putida strains. Moreover, their EPS substrate is easily modified/lost upon resistance development since all isolated infection-resistant strains are also resistant to the formation of halo zones. The in vitro degradation of single-species P. putida biofilms displays a time- and dose-dependent response upon phage inoculation independent of the bacterial strain, phage or age of the pre-grown biofilm. Killing of the associated planktonic cultures occurred in parallel with, but was always more pronounced than the biofilm disintegration. However, the degree of biofilm disintegration and planktonic killing in response to a particular phage is dependent on the bacterial strain involved and/or age of the pre-grown biofilms. No correlation was noted between the phage susceptibility of the biofilm environment and the relative biofilm forming capacity (RBFC) of the specific strain or its susceptibility of its associated planktonic culture. Generally, 24 h after phage inoculation a re-growth of the planktonic culture and biofilm environment was observed which suggest bacterial resistance development to phages. Application of purified tail spike proteins or UV-inactivated phage particles could not disintegrate 24 h pre-grown PpG1 biofilms.The Pseudomonas aeruginosa infecting phages of the phiKMV-like viruses form around their plaques a halo zone which increases in diameter over the course of time. LKA1 only infects strain PAO1 and encodes one tail spike protein Gp49 which forms SDS-resistant trimers and degrades the B-band of the PAO1 lipopolysaccharides. Infection analysis of outer membrane PAO1 mutants indicated that the formation of halo zones upon LKA1 infection is mediated by this tail spike endorhamnosidase activity and is independent of EPS degradation. Thus, LKA1 represents an exception to the current rule of thumb which correlates halo formation and EPS degradation. Despite in vitro antibacterial activity in planktonic cultures of the LKA1 tail spike protein, no in vitro biofilm degradation or killing of its associated planktonic culture was noted. Application of LKA1 phage particles resulted only in a minor biofilm degradation, while a clear time- and dose-dependent killing of the planktonic culture was observed. The other phiKMV-like viruses which contain a tail spike region encoding four proteins, Gp38-41, infect and form halo zones on the alginate-producing strain Pa573. These phages display a fairly broad host spectrum infecting 29% of the P. aeruginosa library. Type IV pili serve as an essential primary receptor, while the AlgC function - involved in alginate biosynthesis is necessary for halo formation. It is expected that Gp38 functions as a versatile adapter which connects the host interacting proteins, Gp40 and 41, to the viral particle. Degradation of the PAO1k and the mucoid Pa573 biofilms by the φKMV-like phage PT-6 appeared to be time- and dose-dependent and decreases with increasing age of the pre-grown biofilms. The planktonic P. aeruginosa cultures were more susceptible for phage infection than their associated biofilms, but both show a re-growth 24 h after inoculation. PT-6-mediated degradation of biofilms formed by twelve susceptible P. aeruginosa strains could be clustered in three groups (no effect, only killing of the associated planktonic culture, combined biofilm degradation and planktonic killing). Some correlation was observed between the RBFC or the clustering of the P. aeruginosa strains and the biofilm/planktonic susceptibility for phage PT-6. No degradation of pre-grown biofilms was observed in the absence of bacterial lysis but with enzymatic active tail spike proteins. The vertical evolution of phages is clearly marked by horizontal transfer of gene(s) (segments) coding for host cell-interacting protein(s) (domains). As such, tail spikes or fibers contain a conserved N-terminal head-binding domain. Horizontal exchange of their C-terminal catalytic domain permits a rapid adaptation of phages to the continually changing host cell surface receptors in the ongoing co-evolutionary arms race with their bacterial hosts. Phages with associated EPS-degrading tail spikes or fibers are able to affect two main constituents of the biofilm environment, the bacterial cells and their EPS molecules. Although, no biofilm degradation is observed in the absence of bacterial lysis, the EPS-degrading tail spikes/fibers are thought to facilitate movement of the phage particles through the biofilm. Probably, the world-wide phage collection contains substantial numbers of phages with not yet recognized EPS-degrading activity. The present data further underscores the need for careful selection of phages for therapeutic purposes. Good phage amplification characteristics in combination with proper EPS depolymerases form a prerequisite, but the final outcome will depend on the environmental conditions and the specific strains involved.status: Publishe
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