bluuebunny/crossref_metadata_2025 huggingface dataset:
Clinical Impact of Preincubation of Blood Cultures at 37°C
Authors: van der Velden Lieven B. , Vos Fidel J. , Mouton Johan W. , Sturm Patrick D.
Published: November 2010

The effect of immediate incubation of blood cultures at 37°C on the turnaround time and the impact of Gram stain results on antimicrobial management were investigated. During a 6-month period, blood cultures collected at the emergency department outside laboratory operating hours were preincubated at 37°C until transportation to the laboratory. Upon the arrival of blood cultures at the laboratory, Gram stains and subcultures were made from all bottles prior to further incubation in the automated system (Bactec 9240). Data from 1 year earlier, when all blood cultures were stored at room temperature, were used for comparison. In the study period, 79 episodes of bacteremia were detected for 75 patients, compared to 70 episodes for 67 patients in the control period. Preincubation of blood cultures at 37°C resulted in a 15-h reduction in the median time to reporting of Gram stain results, from 34 to 19 h (
P
, <0.001). With preincubation, 3 episodes (4%) of bacteremia were not detected by the Bactec 9240 system. Based on the reporting of the Gram stain results, appropriate antimicrobial therapy was initiated for 12% of all patients with positive blood cultures, while for 24% the therapy was streamlined. Thus, immediate incubation of blood cultures reduced the time to reporting of Gram stain results. However, not all episodes of bacteremia were detected by the Bactec 9240 system after preincubation at 37°C. Blood culture results contributed importantly to appropriate antimicrobial management.
Spectrum of Clinically Relevant
Acremonium
Species in the United States
Authors: Perdomo H. , Sutton D. A. , García D. , Fothergill A. W. , Cano J. , Gené J. , Summerbell R. C. , Rinaldi M. G. , Guarro J.
Published: November 2010

Some species in the polyphyletic fungal genus
Acremonium
are important opportunist pathogens. Determining the actual spectrum of species and their incidence in the clinical setting, however, has long been hampered because of the difficulties encountered in phenotypic species-level identification. The goal of this study was to re-identify a large number of clinical isolates morphologically and to confirm the identifications by comparing sequences of the internal transcribed spacer region of the rRNA gene of these isolates to those of type or reference strains of well-known
Acremonium
species. Of the 119 isolates referred to a United States reference laboratory under the name
Acremonium
, only 75 were identified morphologically as belonging to that genus. The remainder (44 isolates) were identified as belonging to other morphologically similar genera. The
Acremonium
clinical isolates were related to species of Hypocreales, Sordariales, and of an
incertae sedis
family of ascomycetes, Plectosphaerellaceae. A total of 50 of the 75
Acremonium
isolates (67%) could be identified by molecular means, the prevalent species being
Acremonium kiliense
(15 isolates),
A. sclerotigenum
-
A. egyptiacum
(11 isolates),
A. implicatum
(7 isolates),
A. persicinum
(7 isolates), and
A. atrogriseum
(4 isolates). One of the most interesting findings of our study was that we identified several species among this large collection of clinical isolates that had not previously been reported from human infections, and we failed to confirm other
Acremonium
species, such as
A. potronii
,
A. recifei
, and
A. strictum
, that had been considered significant. The most common anatomic sites for
Acremonium
isolates were the respiratory tract (41.3%), nails (10.7%), and the eye (9.3%). Antifungal susceptibility testing demonstrated high MICs for all agents tested, except for terbinafine. Since numerous isolates could not be identified, we concluded that the list of opportunistic
Acremonium
species is far from be complete and that a considerable number of additional species will be discovered.
Breakthrough
Hormographiella aspergillata
Infections Arising in Neutropenic Patients Treated Empirically with Caspofungin
Authors: Suarez Felipe , Olivier Gaelle , Garcia-Hermoso Dea , Randriamalala Edouard , Ghez David , Bruneau Julie , Kauffmann-Lacroix Catherine , Bougnoux Marie-Elisabeth , Lortholary Olivier
Published: November 2010

Hormographiella aspergillata
, a filamentous basidiomycete, has rarely been involved in human infections. We describe 2 febrile neutropenic patients who developed a severe pulmonary infection due to
H. aspergillata
while receiving empirical caspofungin therapy for presumed fungal pneumonia. After introduction of liposomal amphotericin B, one patient, who had neutrophil recovery, presented a favorable outcome, while the other, who remained neutropenic throughout the course of infection, died. Resistant fungi, including basidiomycetes, may emerge during empirical treatment with caspofungin in febrile neutropenic patients. A rapid switch to any other potent antifungal should be rapidly considered in case of failure of caspofungin in this setting.
Prevalence and Characterization of Human
Shigella
Infections in Henan Province, China, in 2006
Authors: Xia Shengli , Xu Bianli , Huang Lili , Zhao Jia-Yong , Ran Lu , Zhang Jin , Chen Haomin , Pulsrikarn Chaiwat , Pornruangwong Srirat , Aarestrup Frank M. , Hendriksen Rene S.
Published: November 2010

In 2006, 3,531 fecal samples were collected from patients with diarrhea in Henan Province, China. A total of 467 (13.2%)
Shigella
strains were isolated and serotyped. Seventy-one
Shigella flexneri
strains were characterized by MIC determination, pulsed-field gel electrophoresis (PFGE), and detection of genes encoding cephalosporin resistance. Most infections were caused by
S. flexneri
variant X [IV:(7),8] (27.6%),
S. sonnei
(24.2%), and
S. flexneri
2a (20.8%). However, large regional differences were observed. Significantly higher odds (2.0) of females compared to males were infected with
S. flexneri
2a. Untypeable
S. flexneri
(−:6) isolates were absent among males, as were untypeable
S. flexneri
[I:(7),8] isolates among females. Patient ages ranged from 2 months to 82 years, with 231 subjects (49.7%) <5 years of age. Most of the patients were male (62.1% [
n
= 290]). Infections peaked in July; week 27 with 38 cases (8.1%). All of the 71
S. flexneri
conferred resistance to nalidixic acid; in addition, 21% (
n
= 15) and 79% (
n
= 56) were high- and low-level resistant to ciprofloxacin, respectively. Six
S. flexneri
isolates {serotype 2b [II:7,(8)] and 2b [II:(3),4;7,(8)]} harbored the
bla
CTX-M-14
or
bla
CTX-M-15
gene. A total of 52 unique XbaI PFGE patterns were observed among the 71
S. flexneri
isolates with 11 distinct PFGE clusters. This study revealed a high prevalence of shigellosis with geographical differences in the distribution of serotypes in the distribution of serotypes and also differences in comparisons by gender. A high frequency of resistance, including 100% resistance to ciprofloxacin and resistance to extended-spectrum cephalosporins, was observed. We detected several isolates exhibiting the same PFGE type and MIC profile, indicating multiple undetected outbreaks.
A Twist on Lyme: the Challenge of Diagnosing European Lyme Neuroborreliosis
Authors: Makhani Naila , Morris Shaun K. , Page Andrea V. , Brophy Jason , Lindsay L. Robbin , Banwell Brenda L. , Richardson Susan E.
Published: November 2010

Lyme neuroborreliosis is a tick-borne illness with central and peripheral nervous system manifestations. Clinical features and methods for accurate diagnosis differ across world regions owing to different causative
Borrelia
species. The importance of these distinctions is highlighted by a 12-year-old Canadian girl who acquired Lyme neuroborreliosis in Europe.
HIV-1 Load Comparison Using Four Commercial Real-Time Assays
Authors: Bourlet Thomas , Signori-Schmuck Anne , Roche Laurent , Icard Vinca , Saoudin Henia , Trabaud Mary-Anne , Tardy Jean-Claude , Morand Patrice , Pozzetto Bruno , Ecochard René , André Patrice
Published: November 2010

The HIV-1 RNA viral load is commonly used for the monitoring of disease progression and antiretroviral treatment of HIV-1-infected patients. Since the misestimating of values could lead to inappropriate therapeutical management, the comparative performances, especially the ability to span the genetic diversity of HIV-1, of available automated real-time assays need to be evaluated. We conducted a prospective study with 74 consenting patients enrolled between March 2007 and November 2008. A blood sample was obtained at the time of diagnosis of HIV seropositivity and blindly tested for HIV-1 RNA by at least 4 commercial tests: the Abbott
m
2000 RealTi
m
e HIV-1, bioMérieux NucliSens EasyQ HIV-1, version 1.2 (v1.2), and Cobas AmpliPrep/Cobas TaqMan (CAP/CTM) v1.0 and v2.0 assays. The means of difference were null between CAP/CTM v2.0 and Abbott for CRF02_AG subtypes but positive in favor of CAP/CTM v2.0 for genotype B and negative in favor of NucliSens for all genotypes. The standard deviation (SD) of difference ranged from 0.3 to 0.59, depending on the considered couples of assays. Reliabilities of these four tests, appreciated by the standard deviation of difference between the measurement and the estimated “true” viral load and by the coefficient of reliability, were significantly different (
P
< 10
−4
) among each other. Significant differences were also observed within each group of HIV-1 genotype. The global disparity was higher for CRF02_AG than for B subtypes. This study indicates a risk of viral load misestimating or discrepancies between techniques, depending on the HIV-1 subtype, and speaks in favor of using the same assay for the monitoring of HIV-1-infected patients.
Staphylococcal IgM Enzyme-Linked Immunosorbent Assay for Diagnosis of Periprosthetic Joint Infections
Authors: Artini Marco , Romanò Carlo , Manzoli Lamberto , Scoarughi Gian Luca , Papa Rosanna , Meani Enzo , Drago Lorenzo , Selan Laura
Published: November 2010

Delayed orthopedic joint prosthesis infections (DOJP-Is) due to staphylococci frequently result in prosthetic revision. Specific and noninvasive diagnostic tests are unavailable, and DOJP-Is are commonly diagnosed at advanced stages of disease. An enzyme-linked immunosorbent assay (ELISA) was developed to detect serum antibodies against staphylococcal slime polysaccharide antigens. Using a cutoff of 0.35 ELISA units, the test showed a specificity of 95.1% (95% confidence interval [CI], 85.4 to 98.7%) and a sensitivity of 89.7% (71.5 to 97.3%) on a sample of 90 individuals.
Human Parainfluenza Virus Type 2 L Protein Regions Required for Interaction with Other Viral Proteins and mRNA Capping
Authors: Nishio Machiko , Tsurudome Masato , Garcin Dominique , Komada Hiroshi , Ito Morihiro , Le Mercier Philippe , Nosaka Tetsuya , Kolakofsky Daniel
Published: November 2010

The large RNA polymerase (L) protein of human parainfluenza virus type 2 (hPIV2) binds the nucleocapsid, phosphoprotein, and V protein, as well as itself, and these interactions are essential for transcription and replication of the viral RNA genome. Although all of these interactions were found to be mediated through the domains within the N terminus of L, the C terminus of the L protein was also required for minigenome reporter gene expression. We have identified a highly conserved rubulavirus domain near the C terminus of the L protein that is required for mRNA synthesis but not for genome replication. Remarkably, this region of L shares homology with a conserved region of cellular capping enzymes that binds GTP and forms a lysyl-GMP enzyme intermediate, the first step in the cellular capping reaction. We propose that this conserved region of L also binds GTP (or GDP) to carry out the second step of the unconventional nonsegmented negative-strand virus capping reaction.
The Receptor-Binding Domain of Influenza Virus Hemagglutinin Produced in
Escherichia coli
Folds into Its Native, Immunogenic Structure
Authors: DuBois Rebecca M. , Aguilar-Yañez José Manuel , Mendoza-Ochoa Gonzalo I. , Oropeza-Almazán Yuriana , Schultz-Cherry Stacey , Alvarez Mario Moisés , White Stephen W. , Russell Charles J.
Published: November 2010

The hemagglutinin (HA) surface glycoprotein promotes influenza virus entry and is the key protective antigen in natural immunity and vaccines. The HA protein is a trimeric envelope glycoprotein consisting of a globular receptor-binding domain (HA-RBD) that is inserted into a membrane fusion-mediating stalk domain. Similar to other class I viral fusion proteins, the fusogenic stalk domain spontaneously refolds into its postfusion conformation when expressed in isolation, consistent with this domain being trapped in a metastable conformation. Using X-ray crystallography, we show that the influenza virus HA-RBD refolds spontaneously into its native, immunogenic structure even when expressed in an unglycosylated form in
Escherichia coli
. In the 2.10-Å structure of the HA-RBD, the receptor-binding pocket is intact and its conformational epitopes are preserved. Recombinant HA-RBD is immunogenic and protective in ferrets, and the protein also binds with specificity to sera from influenza virus-infected humans. Overall, the data provide a structural basis for the rapid production of influenza vaccines in
E. coli
. From an evolutionary standpoint, the ability of the HA-RBD to refold spontaneously into its native conformation suggests that influenza virus acquired this domain as an insertion into an ancestral membrane-fusion domain. The insertion of independently folding domains into fusogenic stalk domains may be a common feature of class I viral fusion proteins.
A Dimeric Rep Protein Initiates Replication of a Linear Archaeal Virus Genome: Implications for the Rep Mechanism and Viral Replication
Authors: Oke Muse , Kerou Melina , Liu Huanting , Peng Xu , Garrett Roger A. , Prangishvili David , Naismith James H. , White Malcolm F.
Published: November 2010

The
Rudiviridae
are a family of rod-shaped archaeal viruses with covalently closed, linear double-stranded DNA (dsDNA) genomes. Their replication mechanisms remain obscure, although parallels have been drawn to the
Poxviridae
and other large cytoplasmic eukaryotic viruses. Here we report that a protein encoded in the 34-kbp genome of the rudivirus SIRV1 is a member of the replication initiator (Rep) superfamily of proteins, which initiate rolling-circle replication (RCR) of diverse viruses and plasmids. We show that SIRV Rep nicks the viral hairpin terminus, forming a covalent adduct between an active-site tyrosine and the 5′ end of the DNA, releasing a 3′ DNA end as a primer for DNA synthesis. The enzyme can also catalyze the joining reaction that is necessary to reseal the DNA hairpin and terminate replication. The dimeric structure points to a simple mechanism through which two closely positioned active sites, each with a single tyrosine residue, work in tandem to catalyze DNA nicking and joining. We propose a novel mechanism for rudivirus DNA replication, incorporating the first known example of a Rep protein that is not linked to RCR. The implications for Rep protein function and viral replication are discussed.
Coordinated Leading and Lagging Strand DNA Synthesis by Using the Herpes Simplex Virus 1 Replication Complex and Minicircle DNA Templates
Authors: Stengel Gudrun , Kuchta Robert D.
Published: November 2010

The origin-specific replication of the herpes simplex virus 1 genome requires seven proteins: the helicase-primase (UL5-UL8-UL52), the DNA polymerase (UL30-UL42), the single-strand DNA binding protein (ICP8), and the origin-binding protein (UL9). We reconstituted these proteins, excluding UL9, on synthetic minicircular DNA templates and monitored leading and lagging strand DNA synthesis using the strand-specific incorporation of dTMP and dAMP. Critical features of the assays that led to efficient leading and lagging stand synthesis included high helicase-primase concentrations and a lagging strand template whose sequence resembled that of the viral DNA. Depending on the nature of the minicircle template, the replication complex synthesized leading and lagging strand products at molar ratios varying between 1:1 and 3:1. Lagging strand products (∼0.2 to 0.6 kb) were significantly shorter than leading strand products (∼2 to 10 kb), and conditions that stimulated primer synthesis led to shorter lagging strand products. ICP8 was not essential; however, its presence stimulated DNA synthesis and increased the length of both leading and lagging strand products. Curiously, human DNA polymerase α (p70-p180 or p49-p58-p70-p180), which improves the utilization of RNA primers synthesized by herpesvirus primase on linear DNA templates, had no effect on the replication of the minicircles. The lack of stimulation by polymerase α suggests the existence of a macromolecular assembly that enhances the utilization of RNA primers and may functionally couple leading and lagging strand synthesis. Evidence for functional coupling is further provided by our observations that (i) leading and lagging strand synthesis produce equal amounts of DNA, (ii) leading strand synthesis proceeds faster under conditions that disable primer synthesis on the lagging strand, and (iii) conditions that accelerate helicase-catalyzed DNA unwinding stimulate decoupled leading strand synthesis but not coordinated leading and lagging strand synthesis.
Herpes Simplex Virus Type 1 Helicase-Primase: DNA Binding and Consequent Protein Oligomerization and Primase Activation
Authors: Chen Yan , Bai Ping , Mackay Shannon , Korza George , Carson John H. , Kuchta Robert D. , Weller Sandra K.
Published: November 2010

The heterotrimeric helicase-primase complex of herpes simplex virus type I (HSV-1), consisting of UL5, UL8, and UL52, possesses 5′ to 3′ helicase, single-stranded DNA (ssDNA)-dependent ATPase, primase, and DNA binding activities. In this study we confirm that the UL5-UL8-UL52 complex has higher affinity for forked DNA than for ssDNA and fails to bind to fully annealed double-stranded DNA substrates. In addition, we show that a single-stranded overhang of greater than 6 nucleotides is required for efficient enzyme loading and unwinding. Electrophoretic mobility shift assays and surface plasmon resonance analysis provide additional quantitative information about how the UL5-UL8-UL52 complex associates with the replication fork. Although it has previously been reported that in the absence of DNA and nucleoside triphosphates the UL5-UL8-UL52 complex exists as a monomer in solution, we now present evidence that in the presence of forked DNA and AMP-PNP, higher-order complexes can form. Electrophoretic mobility shift assays reveal two discrete complexes with different mobilities only when helicase-primase is bound to DNA containing a single-stranded region, and surface plasmon resonance analysis confirms larger amounts of the complex bound to forked substrates than to single-overhang substrates. Furthermore, we show that primase activity exhibits a cooperative dependence on protein concentration while ATPase and helicase activities do not. Taken together, these data suggest that the primase activity of the helicase-primase requires formation of a dimer or higher-order structure while ATPase activity does not. Importantly, this provides a simple mechanism for generating a two-polymerase replisome at the replication fork.
Characterization of Bafinivirus Main Protease Autoprocessing Activities
Authors: Ulferts Rachel , Mettenleiter Thomas C. , Ziebuhr John
Published: November 2010

The production of functional nidovirus replication-transcription complexes involves extensive proteolytic processing by virus-encoded proteases. In this study, we characterized the viral main protease (M
pro
) of the type species,
White bream virus
(WBV), of the newly established genus
Bafinivirus
(order
Nidovirales
, family
Coronaviridae
, subfamily
Torovirinae
). Comparative sequence analysis and mutagenesis data confirmed that the WBV M
pro
is a picornavirus 3C-like serine protease that uses a Ser-His-Asp catalytic triad embedded in a predicted two-β-barrel fold, which is extended by a third domain at its C terminus. Bacterially expressed WBV M
pro
autocatalytically released itself from flanking sequences and was able to mediate proteolytic processing in
trans
. Using N-terminal sequencing of autoproteolytic processing products we tentatively identified Gln↓(Ala, Thr) as a substrate consensus sequence. Mutagenesis data provided evidence to suggest that two conserved His and Thr residues are part of the S1 subsite of the enzyme's substrate-binding pocket. Interestingly, we observed two N-proximal and two C-proximal autoprocessing sites in the bacterial expression system. The detection of two major forms of M
pro
, resulting from processing at two different N-proximal and one C-proximal site, in WBV-infected epithelioma papulosum cyprini cells confirmed the biological relevance of the biochemical data obtained in heterologous expression systems. To our knowledge, the use of alternative M
pro
autoprocessing sites has not been described previously for other nidovirus M
pro
domains. The data presented in this study lend further support to our previous conclusion that bafiniviruses represent a distinct group of viruses that significantly diverged from other phylogenetic clusters of the order
Nidovirales
.
Phylogeography of the Spring and Fall Waves of the H1N1/09 Pandemic Influenza Virus in the United States
Authors: Nelson Martha I. , Tan Yi , Ghedin Elodie , Wentworth David E. , St. George Kirsten , Edelman Laurel , Beck Eric T. , Fan Jiang , Lam Tommy Tsan-Yuk , Kumar Swati , Spiro David J. , Simonsen Lone , Viboud Cecile , Holmes Edward C. , Henrickson Kelly J. , Musser James M.
Published: November 2010

Spatial variation in the epidemiological patterns of successive waves of pandemic influenza virus in humans has been documented throughout the 20th century but never understood at a molecular level. However, the unprecedented intensity of sampling and whole-genome sequencing of the H1N1/09 pandemic virus now makes such an approach possible. To determine whether the spring and fall waves of the H1N1/09 influenza pandemic were associated with different epidemiological patterns, we undertook a large-scale phylogeographic analysis of viruses sampled from three localities in the United States. Analysis of genomic and epidemiological data reveals distinct spatial heterogeneities associated with the first pandemic wave, March to July 2009, in Houston, TX, Milwaukee, WI, and New York State. In Houston, no specific H1N1/09 viral lineage dominated during the spring of 2009, a period when little epidemiological activity was observed in Texas. In contrast, major pandemic outbreaks occurred at this time in Milwaukee and New York State, each dominated by a different viral lineage and resulting from strong founder effects. During the second pandemic wave, beginning in August 2009, all three U.S. localities were dominated by a single viral lineage, that which had been dominant in New York during wave 1. Hence, during this second phase of the pandemic, extensive viral migration and mixing diffused the spatially defined population structure that had characterized wave 1, amplifying the one viral lineage that had dominated early on in one of the world's largest international travel centers.
Hepatitis B Virus Regulatory HBx Protein Binds to Adaptor Protein IPS-1 and Inhibits the Activation of Beta Interferon
Authors: Kumar Manish , Jung Sung Yun , Hodgson Amanda J. , Madden Charles R. , Qin Jun , Slagle Betty L.
Published: November 2010

Hepatitis B virus (HBV) encodes the regulatory HBx protein, which is required for virus replication, although its specific role(s) in the replication cycle remains under investigation. An immunoprecipitation/mass spectrometry approach was used to identify four novel HBx binding proteins from the cytoplasmic fraction of HBx transgenic mouse livers. One of these HBx binding partners is beta interferon promoter stimulator 1 (IPS-1), an adaptor protein that plays a critical role in mediating retinoic acid-inducible gene I (RIG-I) signaling, which leads to the activation of beta interferon (IFN-β). The HBx-IPS-1 protein interaction was confirmed in plasmid-transfected HepG2 cells by reciprocal coimmunoprecipitation and Western blotting. We hypothesized that HBx might alter IPS-1 function since proteins of hepatitis C virus and hepatitis A virus similarly bind IPS-1 and target it for inactivation. The effect of HBx on IPS-1-mediated IFN-β signaling was tested in transfected 293T and HepG2 cells, and we show that HBx inhibits double-stranded DNA (dsDNA)-mediated IFN-β activation in a dose-dependent manner when expressed either alone or within the context of HBV replication. However, HBx does not inhibit poly(I:C)-activated IFN-β signaling. These results demonstrate that HBx interferes with the RIG-I pathway of innate immunity. Hepatitis B virus now joins hepatitis C virus and hepatitis A virus in targeting the same innate immune response pathway, presumably as a shared strategy to benefit replication of these viruses in the liver.
The E4orf6/E1B55K E3 Ubiquitin Ligase Complexes of Human Adenoviruses Exhibit Heterogeneity in Composition and Substrate Specificity
Authors: Cheng Chi Ying , Gilson Timra , Dallaire Frédéric , Ketner Gary , Branton Philip E. , Blanchette Paola
Published: November 2010

Although human adenovirus type 5 (Ad5) has been widely studied, relatively little work has been done with other human adenovirus serotypes. The Ad5 E4orf6 and E1B55K proteins form Cul5-based E3 ubiquitin ligase complexes to degrade p53, Mre11, DNA ligase IV, integrin α3, and almost certainly other targets, presumably to optimize the cellular environment for viral replication and perhaps to facilitate persistence or latency. As this complex is essential for the efficient replication of Ad5, we undertook a systematic analysis of the structure and function of corresponding E4orf6/E1B55K complexes from other serotypes to determine the importance of this E3 ligase throughout adenovirus evolution. E4orf6 and E1B55K coding sequences from serotypes representing all subgroups were cloned, and each pair was expressed and analyzed for their capacity to assemble the Cullin-based ligase complex and to degrade substrates following plasmid DNA transfection. The results indicated that all formed Cullin-based E3 ligase complexes but that heterogeneity in both structure and function existed. Whereas Cul5 was present in the complexes of some serotypes, others recruited primarily Cul2, and the Ad16 complex clearly bound both Cul2 and Cul5. There was also heterogeneity in substrate specificity. Whereas all serotypes tested appeared to degrade DNA ligase IV, complexes from some serotypes failed to degrade Mre11, p53, or integrin α3. Thus, a major evolutionary pressure for formation of the adenovirus ligase complex may lie in the degradation of DNA ligase IV; however, it seems possible that the degradation of as-yet-unidentified critical targets or, perhaps even more likely, appropriate combinations of substrates plays a central role for these adenoviruses.
The Presence of a
vpu
Gene and the Lack of Nef-Mediated Downmodulation of T Cell Receptor-CD3 Are Not Always Linked in Primate Lentiviruses
Authors: Schmökel Jan , Sauter Daniel , Schindler Michael , Leendertz Fabian H. , Bailes Elizabeth , Dazza Marie-Christine , Saragosti Sentob , Bibollet-Ruche Frederic , Peeters Martine , Hahn Beatrice H. , Kirchhoff Frank
Published: November 2010

Nef is an accessory protein critical for the ability of human and simian immunodeficiency viruses (HIV and SIV) to replicate efficiently in their respective hosts. Previous analyses of members of 15 different primate lentivirus lineages revealed a link between Nef function and the presence of a
vpu
gene. In particular, Nef proteins of all
vpu
-containing viruses had lost their ability to downmodulate the T cell (TCR-CD3) receptor. Here we examined Nef proteins from eight additional SIV lineages, including SIVgor, SIVwrc, SIVolc, SIVgri, SIVdrl, SIVlho, SIVden, and SIVasc, from western lowland gorillas, western red colobus monkeys, olive colobus monkeys, grivet monkeys, drills, L'Hoest's monkeys, Dent's mona monkeys, and red-tailed monkeys, respectively. We found that except for the
nef
gene of SIVdrl, all of them were efficiently expressed and modulated CD4, major histocompatibility complex class I (MHC-I), CD28, CXCR4, and Ii cell surface expression and/or enhanced viral infectivity and replication. Furthermore, the Nef proteins of SIVgri, SIVlho, SIVwrc, SIVolc, and SIVgor antagonized tetherin. As expected, the Nef protein of SIVgor, which carries
vpu
, failed to downmodulate CD3, whereas those of SIVwrc, SIVgri, SIVlho, and SIVasc, which lack
vpu
, were capable of performing this function. Surprisingly, however, the Nef protein of the
vpu
-containing SIVden strain retained the ability to downmodulate TCR-CD3, whereas that of SIVolc, which does not contain
vpu
, was unable to perform this function. Although the SIVden Vpu is about 20 amino acids shorter than other Vpu proteins, it degrades CD4 and antagonizes tetherin. Our data show that there are exceptions to the link between the presence of a
vpu
gene and
nef
alleles deficient in CD3 modulation, indicating that host properties also affect the selective pressure for Nef-mediated disruption of TCR-CD3 signaling. Our results are also further evidence that tetherin antagonism is a common function of primate lentivirus Nef proteins and that the resistance of human tetherin to Nef represents a relevant barrier to cross-species transmission of SIVs to humans.
Composts containing fluorescent pseudomonads suppress fusarium root and stem rot development on greenhouse cucumber
Authors: Bradley Geoffrey G. , Punja Zamir K.
Published: November 2010

Three composts (Ball, dairy, and greenhouse) were tested for the ability to suppress the development of Fusarium root and stem rot (caused by Fusarium oxysporum f. sp. radicis-cucumerinum) on greenhouse cucumber. Dairy and greenhouse composts significantly reduced disease severity (P = 0.05), while Ball compost had no effect. Assessment of total culturable microbes in the composts showed a positive relationship between disease suppressive ability and total population levels of pseudomonads. In vitro antagonism assays between compost-isolated bacterial strains and the pathogen showed that strains of Pseudomonas aeruginosa exhibited the greatest antagonism. In growth room trials, strains of P. aeruginosa and nonantagonistic Pseudomonas maculicola , plus 2 biocontrol strains of Pseudomonas fluorescens , were tested for their ability to reduce (i) survival of F. oxysporum, (ii) colonization of plants by the pathogen, and (iii) disease severity. Cucumber seedlings grown in compost receiving P. aeruginosa and P. fluorescens had reduced disease severity index scores after 8 weeks compared with control plants without bacteria. Internal stem colonization by F. oxysporum was significantly reduced by P. aeruginosa. The bacteria colonized plant roots at 1.9 × 106 ± 0.73 × 106CFU·(g root tissue)–1and survival was >107 CFU·(g compost)–1after 6 weeks. The locus for 2,4-diacetylphloroglucinol production was detected by Southern blot analysis and confirmed by PCR. The production of the antibiotic 2,4-diacetylphloroglucinol in liquid culture by P. aeruginosa was confirmed by thin layer chromatography. These results demonstrate that composts containing antibiotic-producing P. aeruginosa have the potential to suppress diseases caused by Fusarium species.
Recombinant Porphyromonas gingivalis FimA preproprotein expressed in Escherichia coli is lipidated and the mature or processed recombinant FimA protein forms a short filament in vitro
Authors: Shoji Mikio , Yoshimura Atsutoshi , Yoshioka Hidenobu , Takade Akemi , Takuma Yasuko , Yukitake Hideharu , Naito Mariko , Hara Yoshitaka , Yoshida Shin-Ichi , Nakayama Koji
Published: November 2010

The gram-negative anaerobic bacterium Porphyromonas gingivalis is an etiologically important pathogen for chronic periodontal diseases in adults. Our previous study suggested that the major structural components of both Fim and Mfa fimbriae in this organism are secreted through their lipidated precursors. In this study, we constructed Escherichia coli strains expressing various fimA genes with or without the 5′-terminal DNA region encoding the signal peptide, and we determined whether lipidation of recombinant FimA proteins occurred in E. coli. Lipidation occurred for a recombinant protein from the fimA gene with the 5′-terminal DNA region encoding the signal peptide but not for a recombinant protein from the fimA gene without the signal-peptide-encoding region, as revealed by [3H]palmitic acid labeling experiments. A TLR2-dependent signaling response was induced by the recombinant protein from the fimA gene with the signal-peptide-encoding region but not by a recombinant protein from the fimA gene with the signal-peptide-encoding region that had a base substitution causing an amino acid substitution (C19A). Electron microscopic analysis revealed that recombinant FimA (A-47 – W-383) protein was autopolymerized to form filamentous structures of about 80 nm in length in vitro. The results suggest that FimA protein, a major subunit of Fim fimbriae, is transported to the outer membrane by the lipoprotein sorting system, and a mature or processed FimA protein on the outer membrane is autopolymerized to form Fim fimbriae.
The influence of affinity, efficacy, and slope factor on the estimates obtained by the receptorial responsiveness method (RRM): a computer simulation study
Authors: Grenczer Maria , Pinter Akos , Zsuga Judit , Kemeny-Beke Adam , Juhasz Bela , Szodoray Peter , Tosaki Arpad , Gesztelyi Rudolf
Published: November 2010

The receptorial responsiveness method (RRM) was proposed to characterize changes in the concentration of degradable agonists in the microenvironment of their receptors. The characterization is done by providing concentrations of a stable agonist for the same receptor that is equieffective with the change in concentration to be characterized. RRM is based on the analysis of concentration–effect (E/c) curves reflecting the simultaneous action of the degradable and the stable agonist. In the present study, we investigated whether dissimilar affinity and (or) efficacy of the coacting agonists as well as the steepness of the E/c curves influence the reliability of RRM. E/c curves were simulated based on the operational model and then analyzed with RRM. We found that dissimilarity in affinity of the coacting agonists did not affect the accuracy of RRM estimates. In contrast, accuracy of the estimation depended on the magnitude of the concentration to be assessed, the operational slope factor, and the operational efficacy ratio of the coacting agonists. However, our results suggest that proper choice of a stable agonist for a degradable one can help to ensure reliable results, since information about the change in concentration of a degradable agonist is otherwise difficult to obtain.
Effects of low-dose ketanserin on atherosclerosis in rats and rabbits
Authors: Yu Yong-Sheng , Xie He-Hui , Li Ling , Song Shu-Wei , Han Ping , Cai Guo-Jun , Su Ding-Feng
Published: November 2010

The present study was designed to test the hypothesis that a small dose of ketanserin, which enhances baroreflex activity, prevents the early lesions of atherosclerosis. In experiment 1, baroreflex sensitivity (BRS) was measured in 31 spontaneously hypertensive rats (SHRs) in a conscious state using a computerized blood pressure monitoring system. Four weeks later, the rats were administered vitamin D3and fed a high-cholesterol diet for 8 weeks to induce atherosclerosis. Then their hearts and aortae were removed for pathological examination. A negative correlation was found between BRS and the scores of coronary (r = –0.460, P < 0.01) and aortic atherosclerosis (r = –0.448, P < 0.05) in SHR. In experiment 2, SHRs were divided into 3 groups (n = 10 in each group) and received a dose of ketanserin of 0.3, 1.0, and 3.0 mg/kg (i.g.), respectively. At the smallest dose (0.3 mg/kg), ketanserin did not lower blood pressure but enhanced BRS. In experiment 3, SHRs were administered vitamin D3, fed a high-cholesterol diet, and simultaneously treated with low-dose ketanserin. The atherosclerosis scores of the treatment group were significantly lower than those of the control group (coronary score: 0.90 ± 0.14 vs. 1.76 ± 0.27, P < 0.05; aortic scores: 1.00 ± 0.39 vs. 2.18 ± 0.41, P < 0.05). In experiment 4, male New Zealand White rabbits were fed a high-cholesterol diet and treated with low-dose ketanserin at the same time. The atherosclerosis scores of the treatment group were significantly lower than those of the control group (aortic scores: 0.26 ± 0.20 vs. 0.60 ± 0.31, P < 0.05). In conclusion, the present study demonstrated, for the first time, that low-dose ketanserin prevented the development of atherosclerosis independent of its blood pressure lowering action in SHRs and New Zealand White rabbits at least in part via enhancement of arterial baroreflex function.
Sequential alterations in Akt, GSK3β, and calcineurin signalling in the mouse left ventricle after thoracic aortic constriction
Authors: Prévilon Miresta , Pezet Mylène , Dachez Céline , Mercadier Jean-Jacques , Rouet-Benzineb Patricia
Published: November 2010

Left ventricular hypertrophy (LVH) is an adaptive response to chronic biomechanical stress that generally progresses to maladaptive hypertrophy and heart failure (HF). We studied the activation of protein kinase B (Akt/PKB), glycogen synthase kinase 3 beta (GSK3β), and calcineurin (Cn) at 3, 7, 15, 30, and 60 days following transverse aortic constriction (TAC) in 4-week-old mice. Following TAC, GSK3β inactivation at day 3 was associated with Akt activation, whereas at days 15 and 30, it appeared to be controlled by other kinases. Moderate nonsignificant Cn activation occurred at the early stages, and peak activation at day 30, concomitant with GSK3β inactivation and overt LVH and HF. At the latest stage (day 60), despite further progression of LVH and HF, Cn activation appeared attenuated. Early stages of LVH were associated with Ca2+-handling protein upregulation, whereas major Cn activation, associated with GSK3β inactivation, appeared to engage maladaptive hypertrophy and progression to HF associated with Ca2+-handling protein downregulation.
Regulation of stress-associated scaffold proteins JIP1 and JIP3 on the c-Jun NH2-terminal kinase in ischemia–reperfusion
Authors: Xu Bing , Zhou Yaling , O Karmin , Choy Patrick C. , Pierce Grant N. , Siow Yaw L.
Published: November 2010

Ischemia–reperfusion (IR)-induced cell apoptosis involves the activation of c-Jun NH2-terminal kinase (JNK). The activation of JNK requires the presence of scaffold proteins called JNK-interacting proteins (JIP), which bind several members of a signaling cascade for proper signaling specificity. In this study, the expression of scaffold proteins JIP1 and JIP3 and their roles in the regulation of JNK activity were investigated in simulated IR in a cell model (H9c2). JIP1 protein expression was significantly decreased, whereas JIP3 protein expression was increased in IR H9c2 cells. Adenovirus-induced overexpression of JIP1 reduced IR-induced JNK activity and apoptosis. Conversely, overexpression of JIP3 increased JNK activity and apoptosis following IR. Depletion of endogenous JIP1 by siRNA treatment increased the IR-induced JNK activity, whereas siRNA-mediated depletion of endogenous JIP3 inhibited JNK activity. These results suggest that JIP1 and JIP3 play important roles in the activation of JNK during simulated IR challenge in H9c2 cells.
Coastal niches for terrestrial predators: a stable isotope study
Authors: Mellbrand Kajsa , Hambäck Peter A.
Published: November 2010

The purpose of this study was to identify the use of marine versus terrestrial food items by terrestrial arthropod predators on Baltic Sea shores. The inflow of marine nutrients in the area consists mainly of marine algal detritus and emerging aquatic insects (e.g., chironomids). Diets of coastal arthropods were examined using carbon and nitrogen stable isotope analysis in a two source mixing model. The results suggest that spiders are the terrestrial predators mainly utilizing nutrients and energy of marine origin on Baltic Sea shores, whereas insect predators such as beetles and heteropterans mainly utilize nutrients and energy derived from terrestrial sources, possibly owing to differences in hunting behaviour. That spiders are the predators which benefit the most from the marine inflow suggest that eventual effects of marine subsidies for the coastal ecosystem as a whole are likely mediated by spiders.
FEATURES OF PHOTOCONDUCTIVITY OF Ge AND Si WITH DISORDERED REGIONS
Authors: YEVSEYEV V.
Published: November 2010

Influence of large radiation defects — disordered regions on photoconductivity of semiconductors Ge and Si , compensated as a result of an irradiation with fast neutrons and 1 GeV protons, is investigated. For the first time, the combined role of disordered regions and a large-scale potential relief in photoelectrical properties of Ge and Si , irradiated by high energy particles is defined. The model of photoconductivity that takes into consideration disordered regions as specific sensitizing recombination centers and allows formation of the spatial potential relief is developed.
A STUDY ON MAGNETIC PROPERTIES IN Cu-DOPED ZnO BY THE FIRST PRINCIPLES
Authors: SHI LI-BIN , XU CUI-YAN , YUAN HONG-KUAN
Published: November 2010

Using the first principle method based on density function theory (DFT), we study the electronic and ferromagnetic stability in Cu -doped ZnO . The calculated results based on the local density approximation (LDA) showed that the ferromagnetism (FM) coupling between Cu atoms is more energetically favorable for eight geometrically distinct configurations. In this paper, we also analyze the ferromagnetic properties of Cu -doped ZnO within LDA + U scheme. The dominant ferromagnetic interaction is due to the hybridization between O 2p and Cu 3d. We investigate the effects of oxygen vacancies and nitrogen impurities on FM properties of Cu -doped ZnO . It is obvious that oxygen vacancies and nitrogen impurities are unfavorable in stabilizing the FM of Cu -doped ZnO . In addition, the origin of the FM state of Cu -doped ZnO has also been discussed by analyzing the coupling of Cu 3d levels. Also in this paper, we analyze the strain effect on the FM properties of Cu -doped ZnO .
SUPER SPANNING CONNECTIVITY OF THE FULLY CONNECTED CUBIC NETWORKS
Authors: CHIN CHERNG , CHEN HUAI-CHIH , HSU LIH-HSING , CHIOU SHANG-CHIA , LAI KUO-TUNG
Published: November 2010

A k-containerC(u, v) of G between u and v is a set of k internally disjoint paths between u and v. A k-container C(u,v) of G is a k*-container if it contains all vertices of G. A graph G is k*-connected if there exists a k*-container between any two distinct vertices. The spanning connectivity of G, κ*(G), is defined to be the largest integer k such that G is w*-connected for all 1 ≤ w ≤ k if G is a 1*-connected graph and undefined otherwise. A graph G is super spanning connected if κ* (G) = κ(G). In this paper, we prove that the n-dimensional fully connected cubic network FCCNn is super spanning connected.
Apical and basolateral pools of proteinase-activated receptor-2 direct distinct signaling events in the intestinal epithelium
Authors: Lau Chang , Lytle Christian , Straus Daniel S. , DeFea Kathryn A.
Published: November 2010

Studies suggest that there are two distinct pools of proteinase-activated receptor-2 (PAR2) present in intestinal epithelial cells: an apical pool accessible from the lumen, and a basolateral pool accessible from the interstitial space and blood. Although introduction of PAR2agonists such as 2-furoyl-LIGRL-O-NH2(2fAP) to the intestinal lumen can activate PAR2, the presence of accessible apical PAR2has not been definitively shown. Furthermore, some studies have suggested that basolateral PAR2responses in the intestinal epithelium are mediated indirectly by neuropeptides released from enteric nerve fibers, rather than by intestinal PAR2itself. Here we identified accessible pools of both apical and basolateral PAR2in cultured Caco2-BBe monolayers and in mouse ileum. Activation of basolateral PAR2transiently increased short-circuit current by activating electrogenic Cl−secretion, promoted dephosphorylation of the actin filament-severing protein, cofilin, and activated the transcription factor, AP-1, whereas apical PAR2did not. In contrast, both pools of PAR2activated extracellular signal-regulated kinase 1/2 (ERK1/2) via temporally and mechanistically distinct pathways. Apical PAR2promoted a rapid, biphasic PLCβ/Ca2+/PKC-dependent ERK1/2 activation, resulting in nuclear localization, whereas basolateral PAR2promoted delayed ERK1/2 activation which was predominantly restricted to the cytosol, involving both PLCβ/Ca2+and β-arrestin-dependent pathways. These results suggest that the outcome of PAR2activation is dependent on the specific receptor pool that is activated, allowing for fine-tuning of the physiological responses to different agonists.
The phosphorylation state of MRLC is polyamine dependent in intestinal epithelial cells
Authors: Bavaria Mitulkumar N. , Ray Ramesh M. , Johnson Leonard R.
Published: November 2010

Cell migration is important to the integrity of the gastrointestinal tract for the normal movement of cells from crypt to villi and the healing of wounds. Polyamines are essential to cell migration, mucosal restitution, and, hence, healing. Polyamine depletion by α-difluoromethyl ornithine (DFMO) inhibited migration by decreasing lamellipodia and stress fiber formation and preventing the activation of Rho-GTPases. Polyamine depletion increased the association of the thick F-actin cortex with phosphorylated myosin regulatory light chain (pMRLC). In this study, we determined why MRLC is constitutively phosphorylated as part of the actin cortex. Inhibition of myosin light chain kinase (MLCK) decreased RhoA and Rac1 activities and significantly inhibited migration. Polyamine depletion increased phosphorylation of MRLC (Thr18/Ser19) and stabilized the actin cortex and focal adhesions. The Rho-kinase inhibitor Y27632 increased spreading and migration by decreasing the phosphorylation of MRLC, remodeling focal adhesions, and by activating Rho-GTPases. Thus phosphorylation of MRLC appears to be the rate-limiting step during the migration of IEC-6 cells. In addition, increased localization of RhoA with the actin cortex in polyamine-depleted cells appears to activate Rho-kinase. In the absence of polyamines, activated Rho-kinase phosphorylates myosin phosphatase targeting subunit 1 (MYPT1) at serine-668 leading to its inactivation and preventing the recruitment of phosphatase (protein phosphastase, PP1cδ) to the actomyosin cortex. In this condition, MRLC is constitutively phosphorylated and cycling does not occur. Thus activated myosin binds F-actin stress fibers and prevents focal adhesion turnover, Rho-GTPase activation, and the remodeling of the cytoskeleton required for migration.