bluuebunny/crossref_metadata_2025 huggingface dataset:
Phosphoinositide 3‐kinase mediated signaling in lobster olfactory receptor neurons
Authors: Corey Elizabeth A. , Bobkov Yuriy , Pezier Adeline , Ache Barry W.
Published: February 2010

J. Neurochem. (2010) 113, 341–350.

In vertebrates and some invertebrates, odorant molecules bind to G protein‐coupled receptors on olfactory receptor neurons (ORNs) to initiate signal transduction. Phosphoinositide 3‐kinase (PI3K) activity has been implicated physiologically in olfactory signal transduction, suggesting a potential role for a G protein‐coupled receptor‐activated class I PI3K. Using isoform‐specific antibodies, we identified a protein in the olfactory signal transduction compartment of lobster ORNs that is antigenically similar to mammalian PI3Kγ and cloned a gene for a PI3K with amino acid homology with PI3Kβ. The lobster olfactory PI3K co‐immunoprecipitates with the G protein α and β subunits, and an odorant‐evoked increase in phosphatidylinositol (3,4,5)‐trisphosphate can be detected in the signal transduction compartment of the ORNs. PI3Kγ and β isoform‐specific inhibitors reduce the odorant‐evoked output of lobster ORNs in vivo. Collectively, these findings provide evidence that PI3K is indeed activated by odorant receptors in lobster ORNs and further support the potential involvement of G protein activated PI3K signaling in olfactory transduction.
Reliability and validity of the 2002 and 2006 Chapel Hill expert surveys on party positioning
Authors: HOOGHE LIESBET , BAKKER RYAN , BRIGEVICH ANNA , DE VRIES CATHERINE , EDWARDS ERICA , MARKS GARY , ROVNY JAN , STEENBERGEN MARCO , VACHUDOVA MILADA
Published: February 2010

This research note reports on the 2002 and 2006 Chapel Hill expert surveys (CHES), which measure national party positioning on European integration, ideology, and several European Union (EU) and non‐EU policies. The reliability of expert judgments is examined and the CHES data are cross‐validated with data from the Comparative Manifesto Project, the 2003 Benoit‐Laver expert survey and the 2002 Rohrschneider‐Whitefield survey. The dataset is available on the CHES website.
Open questions in current models of antidepressant action
Authors: Tanti A , Belzung C
Published: February 2010

Research on depression and antidepressant drugs is necessary, as many patients display poor response to therapy. Different symptomatic and pathophysiological features have been proposed as end points of the depressive phenotype and of the antidepressant action, including anhedonia, depressed mood, alterations in morphology and activity of some brain areas (amygdala, nucleus accumbens, hippocampus, prefrontal cortex and cingulate cortex), modifications in the connectivity between brain structures, changes in neurotransmitters (serotonin, noradrenaline, glutamate and neuropeptides), brain plasticity (neurogenesis, neurotrophins) and abnormal function of the hypothalamic‐pituitary adrenal axis. However, few models have been proposed to describe how these end points could induce the depressive phenotype and are involved in the mechanism of action of antidepressants. Here we propose a connectionist‐inspired network of depression and antidepressant action, in which the different aetiological factors participating in the release of a depressive episode are represented by input nodes, the different symptomatic as well as pathophysiological end points are represented by an intermediate layer, and the onset of depression or of comorbid disease is represented by the output node. The occurrence of depression and the mechanism of the antidepressant action thus depend upon the weight of the interactions between the different end points, none of them beingper secrucial to the onset of a depressive phenotype or to the antidepressant action. This model is heuristic to draw future lines of research concerning new antidepressant therapies, designing new animal models of depression and for a better understanding of the depressive pathology and of its comorbid pathology such as anxiety disorders.
Mechanisms involved in the antinociception induced by systemic administration of guanosine in mice
Authors: Schmidt AP , Böhmer AE , Schallenberger C , Antunes C , Tavares RG , Wofchuk ST , Elisabetsky E , Souza DO
Published: February 2010

Background and purpose: It is well known that adenine‐based purines exert multiple effects on pain transmission. However, less attention has been given to the potential effects of guanine‐based purines on pain transmission. The aim of this study was to investigate the effects of intraperitoneal (i.p.) and oral (p.o.) administration of guanosine on mice pain models. Additionally, investigation into the mechanisms of action of guanosine, its potential toxicity and cerebrospinal fluid (CSF) purine levels were also assessed.

Experimental approach: Mice received an i.p. or p.o. administration of vehicle (0.1 mM NaOH) or guanosine (up to 240 mg·kg−1) and were evaluated in several pain models.

Key results: Guanosine produced dose‐dependent antinociceptive effects in the hot‐plate, glutamate, capsaicin, formalin and acetic acid models, but it was ineffective in the tail‐flick test. Additionally, guanosine produced a significant inhibition of biting behaviour induced by i.t. injection of glutamate, AMPA, kainate and trans‐ACPD, but not against NMDA, substance P or capsaicin. The antinociceptive effects of guanosine were prevented by selective and non‐selective adenosine receptor antagonists. Systemic administration of guanosine (120 mg·kg−1) induced an approximately sevenfold increase on CSF guanosine levels. Guanosine prevented the increase on spinal cord glutamate uptake induced by intraplantar capsaicin.

Conclusions and implications: This study provides new evidence on the mechanism of action of the antinociceptive effects after systemic administration of guanosine. These effects seem to be related to the modulation of adenosine A1and A2Areceptors and non‐NMDA glutamate receptors.
Ligand‐directed signalling at β‐adrenoceptors
Authors: Evans Bronwyn A. , Sato Masaaki , Sarwar Mohsin , Hutchinson Dana S. , Summers Roger J.
Published: February 2010

β‐Adrenoceptors (ARs) classically mediate responses to the endogenous ligands adrenaline and noradrenaline by coupling to Gsα and stimulating cAMP production; however, drugs designed as β‐AR agonists or antagonists can activate alternative cell signalling pathways, with the potential to influence clinical efficacy. Furthermore, drugs acting at β‐ARs have differential capacity for pathway activation, described as stimulus trafficking, biased agonism, functional selectivity or ligand‐directed signalling. These terms refer to responses where drug A has higher efficacy than drug B for one signalling pathway, but a lower efficacy than drug B for a second pathway. The accepted explanation for such responses is that drugs A and B have the capacity to induce or stabilize distinct active conformations of the receptor that in turn display altered coupling efficiency to different effectors. This is consistent with biophysical studies showing that drugs can indeed promote distinct conformational states. Agonists acting at β‐ARs display ligand‐directed signalling, but many drugs acting as cAMP antagonists are also able to activate signalling pathways central to cell survival and proliferation or cell death. The observed complexity of drug activity at β‐ARs, prototypical G protein‐coupled receptors, necessitates rethinking of the approaches used for screening and characterization of novel therapeutic agents. Most studies of ligand‐directed signalling employ recombinant cell systems with high receptor abundance. While such systems are valid for examining upstream signalling events, such as receptor conformational changes and G protein activation, they are less robust when comparing downstream signalling outputs as these are likely to be affected by complex pathway interactions.

This article is part of a themed section on Molecular Pharmacology of GPCR. To view the editorial for this themed section visithttp://dx.doi.org/10.1111/j.1476‐5381.2010.00695.x
Ligand binding assays at equilibrium: validation and interpretation
Authors: Hulme Edward C. , Trevethick Mike A.
Published: February 2010

The focus of this review paper is factors affecting data interpretation in ligand binding assays under equilibrium conditions. Protocols for determiningKd(the equilibrium dissociation constant) andKdA(the equilibrium inhibitor constant) for receptor ligands are discussed. The basic theory describing the interaction of a radiotracer and an unlabelled competitor ligand with a receptor is developed. Inappropriate experimental design may result in ligand depletion and non‐attainment of equilibrium, distorting the calculation ofKdandKdA. Strategies, both theoretical and practical, will be given to avoid and correct such errors, thus leading to the determination of reliable values for these constants. In determiningKdAfrom competition binding studies, two additional concepts are discussed. First, the necessity to measure an adequate specific binding signal from the bound radiotracer ligand limits the range of affinity constants that can be measured: a particular set of assay conditions may lead to an upper limit on the apparent affinity of unlabelled ligands. Second, an extension of the basic assay methodology can indicate whether the interaction between the tracer and a test ligand is mediated by a competitive or an allosteric mechanism. Finally, the review ends with a discussion of two factors that are often overlooked: buffer composition and the temperature at which the assay is conducted, and the impact these can have on affinity measurements and the understanding of drug interactions.British Journal of Pharmacology(2010)161, 1219–1237; doi:10.1111/j.1476‐5381.2009.00604.x; published online 2 February 2010

This article is part of a themed section on Analytical Receptor Pharmacology in Drug Discovery. To view the other articles in this section visithttp://dx.doi.org/10.1111/bph.2010.161.issue‐6
Comparative Expression of Proteins in Left and Right Atrial Appendages From Patients With Mitral Valve Disease at Sinus Rhythm and Atrial Fibrillation
Authors: MODREGO JAVIER , MAROTO LUIS , TAMARGO JUAN , AZCONA LUIS , MATEOS‐CÁCERES PETRA , SEGURA ANTONIO , MORENO‐HERRERO ROSARIO , PÉREZ‐CASTELLANOS NICASIO , DELPÓN EVA , PÉREZ‐VILLACASTÍN JULIÁN , RODRÍGUEZ ENRIQUE , MACAYA CARLOS , LÓPEZ‐FARRÉ ANTONIO J.
Published: February 2010

Proteomics and Atrial Appendages. Introduction: The objective was to compare by proteomics the expression of proteins associated with the cytoskeleton, energetic metabolism, and cardiac cytoprotection between left atrial appendages (LAA) and right atrial appendages (RAA) obtained from patients with mitral valve disease both in sinus rhythm (SR, n = 6) and in permanent atrial fibrillation (AF, n = 11).

Methods and Results:
Samples from RAA and LAA were obtained from the same patient. Proteins were separated in 2‐dimensional electrophoresis and identified by mass spectrometry. LAA from SR patients upexpressed α‐actin isotype 1 and desmin isotypes 3 and 5 with respect to RAA. In LAA from AF patients were upexpressed cardiac α‐actin isotypes 1 and 2, tropomyosin α‐ and β‐chains, and myosin light chain embryonic muscle/atrial isoform with respect to LAA from SR patients. In RAA from AF patients also upexpressed different cytoskeleton associated proteins with respect to RAA from SR patients. Different energetic metabolism‐associated proteins were upexpressed in LAA and RAA from AF with respect those from SR patients. In AF patients, the expression of proteins associated with cardiac cytoprotection such as gluthatione‐S‐transferase, heat shock protein (Hsp) 27, and different Hsp60 isotypes, were higher in RAA but not in LAA with respect to the corresponding appendages in SR patients.

Conclusions:
For each individual patient RAA and LAA showed a similar level of proteins expressed associated with cytoskeleton, energetic metabolism, and cardiac cytoprotection. There were more differences in the level of proteins associated with the above‐mentioned mechanisms between the atrial appendages from AF with respect to SR patients, which may open new targets for drugs. (J Cardiovasc Electrophysiol, Vol. 21, pp. 859‐868, August 2010)
Intracoronary Beta‐Radiation Therapy for In‐stent Restenosis: Long‐Term Success Rate and Prediction of Failure
Authors: VLACHOJANNIS GEORGIOS J. , FICHTLSCHERER STEPHAN , SPYRIDOPOULOS IOAKIM , AUCH‐SCHWELK WOLFGANG , SCHOPOHL BERND , ZEIHER ANDREAS M. , SCHÄCHINGER VOLKER
Published: February 2010

Objective: The following retrospective observational study assesses the long‐term results of intracoronary beta‐radiation therapy for patients with in‐stent restenosis.

Background: Beta‐radiation has been used to treat patients with coronary in‐stent restenosis. However, long‐term clinical success using this technique has not at this time been established.

Methods: Two‐hundred and thirteen consecutive patients received intracoronary brachytherapy (noncentered beta‐emitter, Novoste BetaCath™) for in‐stent restenosis and were followed up over a period of 39.1 ± 18.4 months. The combined end‐point was defined as a major adverse clinical event (MACE)
and comprised mortality, acute myocardial infarction, or target vessel revascularization (TVR).

Results: MACE occurred in 110 patients (51.6%): death in 27 patients (12.7%), acute myocardial infarction in 8 patients (3.8%), TVR in 90 patients (42.3%). TVR comprised percutaneous coronary reinterventions in 76 patients (35.7%) and coronary bypass surgery in 24 patients (11.3%). Secondary end‐point was determined as target vessel failure and occurred in 93 patients (43.7%). Of note, the frequency of at least two previous target lesion interventions as well as impairment of left ventricular function was associated with reduced success rate, whereas other clinical parameters did not indicate outcome after treatment with intracoronary radiation therapy.

Conclusion: During the mean, a period of 3 years, more than half of the patients receiving intracoronary radiation therapy reached primary end‐point, representing, in the main, TVR. During this period a mortality rate of nearly 13% was documented. These results signify a delayed, though continued, restenotic process after index procedure. (J Interven Cardiol 2010;23:60–65)
Whole brain radiation alone produces favourable outcomes for AIDS‐related primary central nervous system lymphoma in the HAART era
Authors: Nagai Hirokazu , Odawara Takashi , Ajisawa Atsushi , Hagiwara Shotaro , Watanabe Tomoyuki , Uehira Tomoko , Uchiumi Hideki , Yotsumoto Mihoko , Miyakawa Toshikazu , Watanabe Akira , Kambe Toshiyuki , Konishi Mitsuru , Saito Seiji , Takahama Soichiro , Tateyama Masao , Okada Seiji
Published: February 2010

Primary central nervous system lymphoma (PCNSL) related to acquired immunodeficiency syndrome (AIDS) is a lethal disorder, but the recent application of highly active antiretroviral therapy (HAART) has significantly improved prognosis. This retrospective cohort study of AIDS‐related PCNSL examined the actual clinical outcomes and prognostic variables affecting overall survival (OS) in the HAART era. Twenty‐three newly diagnosed AIDS‐related PCNSL at 12 regional centre hospitals for HIV/AIDS in Japan between 2002 and 2008 were consecutively enrolled. The estimated 3‐yr OS rate of the entire cohort was 64% (95%CI, 41.0–80.3%). Whole brain radiation therapy (WBRT) had an independent positive impact on survival (WBRT ≥30 Gy vs. others, P = 0.02). Nine of 10 patients with a good performance status (PS) (0–2) remained alive with complete response, whereas 10 (77%) of 13 of those with a poor PS (3–4) died mostly after a short period. The estimated 3‐yr OS rate of the groups with a good and poor PS was 100% and 38% (95%CI, 14–63%), respectively (P = 0.01). Leukoencephalopathy (grade ≥ 2) developed in 21% of those that survived more than 12 months after radiation. The patients receiving a curative intent radiation dose (≥30 Gy) of WBRT achieved prolonged survival while maintaining a good quality of life in the HAART era, especially among patients with a favourable PS.
White matter changes in extremely preterm infants, a population‐based diffusion tensor imaging study
Authors: Skiöld Béatrice , Horsch Sandra , Hallberg Boubou , Engström Mathias , Nagy Zoltan , Mosskin Mikael , Blennow Mats , Ådén Ulrika
Published: February 2010

Aim:  To investigate cerebral white matter (WM) abnormalities (J Pediatr 2003; 143: 171) and diffuse and excessive high signal intensities (DEHSI), (J Pediatr 1999; 135: 351) in a cohort of extremely preterm infants born in Stockholm during a 3‐year period, using magnetic resonance imaging (MRI) and diffusion tensor imaging (DTI).

Methods:  MRI at term‐equivalent age was performed in 109 infants and DTI data were acquired in 54 infants. Survival rate in the entire cohort was 67%. Sixteen term‐born healthy control infants were scanned for comparison.

Results:  No or mild WM abnormalities were seen in 86% of infants and 14% had moderate or severe WM abnormalities. DEHSI were seen in infants with all grades of white matter abnormalities and were present in 56% of infants. In the WM at the level of centrum semiovale, infants with any WM abnormalities or DEHSI had lower Fractional Anisotropy and higher Apparent Diffusion Coefficient compared with control infants. No significant differences in diffusion were seen in infants without DEHSI compared with the controls in this region. Compared with controls, the preterm infants had significantly altered diffusion in the corpus callosum.

Conclusion:  Only 14% of the extremely preterm infants had moderate or severe WM abnormalities on MRI. However, the incidence of DEHSI was high. In the DEHSI regions, changes in diffusion parameters were detected, indicating altered WM organization.
A direct limit on the turbulent velocity of the intracluster medium in the core of Abell 1835 from XMM-Newton
Authors: Sanders J. S. , Fabian A. C. , Smith R. K. , Peterson J. R.
Published: February 2010

We examine deep XMM–Newton Reflection Grating Spectrometer observations of the X-ray luminous galaxy cluster A1835. For the first time in a galaxy cluster, we place direct limits on turbulent broadening of the emission lines. This is possible because the coolest X-ray emitting gas in the cluster, which is responsible for the lines, occupies a small region within the core. The most conservative determination of the 90 per cent upper limit on line-of-sight, non-thermal, velocity broadening is , measured from the emission lines originating within 30 kpc radius. The ratio of turbulent to thermal energy density in the core is therefore less than 13 per cent. There are no emission lines in the spectrum showing evidence for gas below ∼3.5 keV. We examine the quantity of gas as a function of temperature and place a limit of (90 per cent) for gas cooling radiatively below 3.85 keV.
An Inside‐Outsider or an Outside‐Insider? The Republican Primary Campaign of Ron Paul from a Third‐Party Perspective
Authors: CHAMBERLAIN ADAM
Published: February 2010

The 2007/2008 Republican presidential primary run for Ron Paul was a relatively unique candidacy, as he was a former third‐party presidential candidate for the Libertarian Party in 1988. In this article, I argue that his recent run as a Republican needs to be studied as a third‐party movement within a major party. To do so, I conduct a state‐level analysis of support for Paul using a dependent variable measuring the number of online Paul donors. The results indicate that the strongest explanation for Paul support is a state's vote for Paul in 1988. This finding shows that a third‐party campaign can help a politician's national‐level career rather than serving merely as an outlet for popular discontent. It also shows that candidate‐centered support can carry over into a major‐party campaign, even if almost 20 years has elapsed and a candidate's third‐party vote percentage was quite low.

La primaria presidencial republicana de 2007/2008 donde se postuló Ron Paul fue una candidatura relativamente única, ya que en 1988 él fue un candidato presidencial de un tercer partido conocido como Partido Liberal. En este articulo sostengo que su reciente postulación como Republicano necesita ser estudiada como un movimiento de un tercer partido dentro de un partido mayoritario. Para realizarlo, realizo un análisis a un nivel estatal del apoyo a Paul utilizando una variable dependiente que mide el número de los donadores de Paul en línea. Los resultados indican que la explicación más concluyente del actual apoyo a Paul es el voto estatal otorgado al candidato en 1988. Este hallazgo demuestra que una campaña de un tercer partido puede ayudar a la carrera de un político a nivel nacional más que simplemente servir como una salida del descontento popular. Así mismo, demuestra que el apoyo otorgado al candidato puede transferirse a una campaña de partido mayor, aun después de veinte años y a pesar de que el porcentaje de voto obtenido por ese candidato en ese entonces haya sido bajísimo.
Cardiac basal metabolism: energetic cost of calcium withdrawal in the adult rat heart
Authors: Bonazzola P. , Takara D.
Published: February 2010

Aim:  Cardiac basal metabolism upon extracellular calcium removal and its relationship with intracellular sodium and calcium homeostasis was evaluated.

Methods:  A mechano‐calorimetric technique was used that allowed the simultaneous and continuous measurement of both heat rate and resting pressure in arterially perfused quiescent adult rat hearts. Using pharmacological tools, the possible underlying mechanisms related to sodium and calcium movements were investigated.

Results:  Resting heat rate (expressed in mW g−1dry wt) increased upon calcium withdrawal (+4.4 ± 0.2). This response was: (1) unaffected by the presence of tetrodotoxin (+4.3 ± 0.6), (2) fully blocked by both, the decrease in extracellular sodium concentration and the increase in extracellular magnesium concentration, (3) partially blocked by the presence of either nifedipine (+2.8 ± 0.4), KB‐R7943 (KBR; +2.5 ± 0.2), clonazepam (CLO; +3.1 ± 0.3) or EGTA (+1.9 ± 0.3). The steady heat rate under Ca2+‐free conditions was partially reduced by the addition of Ru360 (−1.1 ± 0.2) but not CLO in the presence of EGTA, KBR or Ru360.

Conclusion:  Energy expenditure for resting state maintenance upon calcium withdrawal depends on the intracellular rise in both sodium and calcium. Our data are consistent with a mitochondrial Ca2+ cycling, not detectable under normal calcium diastolic levels. The experimental condition here analysed, partially simulates findings reported under certain pathological situations including heart failure in which mildly increased levels of both diastolic sodium and calcium have also been found. Therefore, under such pathological conditions, hearts should distract chemical energy to fuel processes associated with sodium and calcium handling, making more expensive the maintenance of their functions.
The care of older people with dementia in acute hospitals
Authors: Cowdell Fiona
Published: February 2010

cowdell f. (2010) The care of older people with dementia in acute hospitals. International Journal of Older People Nursing 5, 83–92.

Aim.  To explore the experiences of patients and nursing staff of the care received by older people with dementia in acute hospitals.

Background.  The prevalence of dementia is steadily increasing as is the number of people with the condition requiring acute hospital care. Significant concerns about the quality of this care have been raised. There is a paucity of knowledge about the views of such care from the perspectives of people with dementia and nurses.

Method.  An ethnographic approach was used and data were collected thorough observation and interviews in one acute hospital in the United Kingdom.

Findings.  Findings suggest that care for older people with dementia in acute hospitals is not always optimum although there are clear exceptions. Generally, people with dementia found the delivery of care and the experience of being in hospital distressing as they did not know what was happening and they were often ignored. Nurses strive to give good care but do not always achieve this.

Conclusion.  Bourdieu’s Model of Practice assists in explaining why care is as it is. There is a clear need to improve current practice.

Relevance to clinical practice.  It is imperative that innovative methods of developing practice are implemented and evaluated. Education alone will not lead to sustained changes in practice. Further research into this subject needs to be undertaken.
On complete interpolating sequences and sampling expansions
Authors: Smith Kevin
Published: February 2010

Complete interpolating sequences for L2(−π,π) are considered under the condition that the real parts of the sequence are a subsequence of the scaled integers xℤ, x>0. It is found that this condition leads to very specific and restrictive conditions on the existence and structure of complete interpolating sequences for L2(−π,π). Further general results in the case of bunched sampling of Bernstein functions are also given.
Platelet Microparticles Drive Inflammatory Arthritis
Authors: Mueller Kristen
Published: February 2010

Microparticles released by platelets contribute to inflammation underlying rheumatoid arthritis.
Keeping the Host Live
Authors: Gough Nancy R.
Published: February 2010

Parasitic worms secrete proteases that when internalized suppress the immune response to promote an anti-inflammatory environment.
Gut Check
Authors: Kiberstis Paula A.
Published: February 2010

Ionizing radiation destroys gastrointestinal epithelial cells by a mechanism that appears to be independent of apoptosis.
BMPs: From Bone to Body Morphogenetic ProteinsA report on the First International BMP Workshop: “Modern Trends in BMP Signaling,” Berlin, Germany, 6 to 9 September 2009.
Authors: Obradovic Wagner Darja , Sieber Christina , Bhushan Raghu , Börgermann Jan H. , Graf Daniel , Knaus Petra
Published: February 2010

Discussion at a meeting in Berlin, Germany, showed that BMPs have essential functions in organs and tissues besides bone.
Genetic and Biochemical Characterization of TRU-1, the Endogenous Class C β-Lactamase fromAeromonas enteropelogenes
Authors: De Luca Filomena , Giraud-Morin Chantal , Rossolini Gian Maria , Docquier Jean-Denis , Fosse Thierry
Published: February 2010

Aeromonas enteropelogenes(formerlyA. tructi) was described to be an ampicillin-susceptible and cephalothin-resistantAeromonasspecies, which suggests the production of a cephalosporinase. Strain ATCC 49803 was susceptible to amoxicillin, cefotaxime, and imipenem but resistant to cefazolin (MICs of 2, 0.032, 0.125, and >256 μg/ml, respectively) and produced an inducible β-lactamase. Cefotaxime-resistant mutants (MIC, 32 μg/ml) that showed constitutive β-lactamase production could be selectedin vitro. The gene coding for the cephalosporinase ofA. enteropelogenesATCC 49803 was cloned, and its biochemical properties were investigated.Escherichia colitransformants showing resistance to various β-lactams carried a 3.5-kb plasmid insert whose sequence revealed a 1,146-bp open reading frame (ORF) encoding a class C β-lactamase, named TRU-1, showing the highest identity scores withA. punctataCAV-1 (75%),A. salmonicidaAmpC (75%), andA. hydrophilaCepH (71%). TheblaTRU-1locus includes open reading frames (ORFs) showing significant homology with genes found in the genomes of otherAeromonasspecies, although it exhibits a different organization, as reflected by the presence of additional ORFs located downstream of the β-lactamase gene in theA. hydrophilaandA. salmonicidagenomes. Specific PCR assays were negative forcphA-like andblaOXA-12-like genes in threeA. enteropelogenesATCC strains. Purified TRU-1 showed a broad substrate profile, efficiently hydrolyzing benzylpenicillin, cephalothin, cefoxitin, and, although with significantly lower turnover rates, oxyiminocephalosporins. Cephaloridine and cefepime were poorly recognized by the enzyme, as reflected by the highKmvalues observed with these substrates. Thus far,A. enteropelogenesrepresents the only known example of anAeromonasspecies that produces only one β-lactamase belonging to molecular class C.
Inhibition of HIV-1 Replication by a Bis-Thiadiazolbenzene-1,2-Diamine That Chelates Zinc Ions from Retroviral Nucleocapsid Zinc Fingers
Authors: Pannecouque Christophe , Szafarowicz Beata , Volkova Natalia , Bakulev Vasiliy , Dehaen Wim , Mély Yves , Daelemans Dirk
Published: February 2010

The human immunodeficiency virus type 1 (HIV-1) nucleocapsid p7 (NCp7) protein holds two highly conserved “CCHC” zinc finger domains that are required for several phases of viral replication. Basic residues flank the zinc fingers, and both determinants are required for high-affinity binding to RNA. Several compounds were previously found to target NCp7 by reacting with the sulfhydryl group of cysteine residues from the zinc fingers. Here, we have identified an
N
,
N
′-bis(1,2,3-thiadiazol-5-yl)benzene-1,2-diamine (NV038) that efficiently blocks the replication of a wide spectrum of HIV-1, HIV-2, and simian immunodeficiency virus (SIV) strains. Time-of-addition experiments indicate that NV038 interferes with a step of the viral replication cycle following the viral entry but preceding or coinciding with the early reverse transcription reaction, pointing toward an interaction with the nucleocapsid protein p7. In fact,
in vitro
, NV038 efficiently depletes zinc from NCp7, which is paralleled by the inhibition of the NCp7-induced destabilization of cTAR (complementary DNA sequence of TAR). A chemical model suggests that the two carbonyl oxygens of the esters in this compound are involved in the chelation of the Zn
2+
ion. This compound thus acts via a different mechanism than the previously reported zinc ejectors, as its structural features do not allow an acyl transfer to Cys or a thiol-disulfide interchange. This new lead and the mechanistic study presented provide insight into the design of a future generation of anti-NCp7 compounds.
Pathogenicity Islands PAPI-1 and PAPI-2 Contribute Individually and Synergistically to the Virulence of
Pseudomonas aeruginosa
Strain PA14
Authors: Harrison Ewan M. , Carter Melissa E. K. , Luck Shelley , Ou Hong-Yu , He Xinyi , Deng Zixin , O'Callaghan Chris , Kadioglu Aras , Rajakumar Kumar
Published: February 2010

Pseudomonas aeruginosa
is a leading cause of hospital-acquired pneumonia and severe chronic lung infections in cystic fibrosis patients. The reference strains PA14 and PAO1 have been studied extensively, revealing that PA14 is more virulent than PAO1 in diverse infection models. Among other factors, this may be due to two pathogenicity islands, PAPI-1 and PAPI-2, both present in PA14 but not in PAO1. We compared the global contributions to virulence of PAPI-1 and PAPI-2, rather than that of individual island-borne genes, using murine models of acute pneumonia and bacteremia. Three isogenic island-minus mutants (PAPI-1-minus, PAPI-2-minus, and PAPI-1-minus, PAPI-2-minus mutants) were compared with the wild-type parent strain PA14 and with PAO1. Our results showed that both islands contributed significantly to the virulence of PA14 in acute pneumonia and bacteremia models. However, in contrast to the results for the bacteremia model, where each island was found to contribute individually, loss of the 108-kb PAPI-1 island alone was insufficient to measurably attenuate the mutant in the acute pneumonia model. Nevertheless, the double mutant was substantially more attenuated, and exhibited a lesser degree of virulence, than even PAO1 in the acute pneumonia model. In particular, its ability to disseminate from the lungs to the bloodstream was markedly inhibited. We conclude that both PAPI-1 and PAPI-2 contribute directly and synergistically in a major way to the virulence of PA14, and we suggest that analysis of island-minus strains may be a more appropriate way than individual gene knockouts to assess the contributions to virulence of large, horizontally acquired segments of DNA.
Acute Immune Response toMycobacterium massiliensein C57BL/6 and BALB/c Mice
Authors: Martins de Sousa Eduardo , Bonfim de Bortoli Fernando , Amaral Eduardo Pinheiro , Batista Aline Carvalho , Liberman Kipnis Thereza , Marques Cardoso Alessandra , Kipnis André , Junqueira-Kipnis Ana Paula
Published: February 2010

Mycobacterium massilienseis an environmental opportunistic pathogen that has been associated with soft tissue infection after minor surgery. We studied the acute immune response of C57BL/6 and BALB/c mice infected intravenously with 106CFU of anM. massiliensestrain isolated from a nosocomial infection in Brazil. The results presented here show thatM. massilienseis virulent and pathogenic to both C57BL/6 and BALB/c mice, inducing a granulomatous inflammatory reaction that involves the activation of macrophages, dendritic cells, and natural killer cells induced by gamma interferon and interleukin-17 (IL-17) in C57BL/6 mice and by IL-12 in BALB/c mice.
Toll-Like Receptor 3 Agonist Protection against ExperimentalFrancisella tularensisRespiratory Tract Infection
Authors: Pyles Richard B. , Jezek G. Eric , Eaves-Pyles Tonyia D.
Published: February 2010

We investigated whether Toll-like receptor 3 (TLR3) stimulation would protect the host from inhaledFrancisella tularensis. TLR3 is expressed by respiratory epithelial cells and macrophages and can be activated by a synthetic double-stranded RNA ligand called polyinosine-polycytosine [poly(I:C)]. Thus, we evaluated poly(I:C) as a novel treatment against inhaledF. tularensis.In vivo, BALB/c mice intranasally (i.n.) treated with poly(I:C) (100 μg/mouse) 1 h before or after Schu 4 or LVS (100 CFU) i.n. challenge showed that poly(I:C) treatment significantly reduced bacterial load in the lungs (P< 0.05). Bronchoalveolar lavage from poly(I:C)-treated mice alone or combined withF. tularensisinfection significantly increased cytokine secretion and enhanced neutrophil influx to lung tissues. Poly(I:C) responses were transient but significantly prolonged the survival of treated mice after i.n.F. tularensischallenge relative to mock treated animals. This prolonged survival providing a longer window for initiation of levofloxacin (LEVO) treatment (40 mg/kg). Animals treated with poly(I:C), challenged withF. tularensis, and then treated with LEVO 5 days later had 100% survival relative to 0% survival in animals receiving LEVO alone. Mechanistically, poly(I:C) given to human monocyte-derived macrophages before or after Schu 4 or LVS challenge (multiplicity of infection, 20:1) had significantly reduced intracellular bacterial replication (P< 0.05). These data suggest that poly(I:C) may represent a potential therapeutic agent against inhaledF. tularensisthat prolongs survival and the opportunity to initiate standard antibiotic therapy (i.e., LEVO).
Contribution of Each of Four Superantigens toStreptococcus equi-Induced Mitogenicity, Gamma Interferon Synthesis, and Immunity
Authors: Paillot Romain , Robinson Carl , Steward Karen , Wright Nicola , Jourdan Thibaud , Butcher Nicola , Heather Zoe , Waller Andrew S.
Published: February 2010

Streptococcus equiis the causative agent of strangles, the most frequently diagnosed infectious disease of horses worldwide. The disease is characterized by abscessation and swelling of the lymph nodes of the head and neck, which can literally strangle the horse to death.S. equiproduces four recently acquired phage-associated bacterial superantigens (sAgs; SeeH, SeeI, SeeL, and SeeM) that share homology with the mitogenic toxins ofStreptococcus pyogenes. The aim of this study was to characterize the contribution of each of theseS. equisAgs to mitogenic activityin vitroand quantify the sAg-neutralizing capacity of sera from naturally infected horses in order to better understand their role in pathogenicity. Each of the sAgs was successfully cloned, and soluble proteins were produced inEscherichia coli. SeeI, SeeL, and SeeM induced a dose-dependent proliferative response in equine CD4 T lymphocytes and synthesis of gamma interferon (IFN-γ). SeeH did not stimulate equine peripheral blood mononuclear cells (PBMC) but induced proliferation of asinine PBMC. Allelic replacement mutants ofS. equistrain 4047 with sequential deletion of the superantigen genes were generated. Deletion ofseeI, seeL, andseeMcompletely abrogated the mitogenic activity and synthesis of IFN-γ, in equine PBMC, of the strain 4047 culture supernatant. Sera from naturally infected convalescent horses had only limited sAg-neutralizing activities. We propose thatS. equisAgs play an important role inS. equipathogenicity by stimulating an overzealous and inappropriate Th1 response that may interfere with the development of an effective immune response.
Virulence Plasmid Harbored by UropathogenicEscherichia coliFunctions in Acute Stages of Pathogenesis
Authors: Cusumano Corinne K. , Hung Chia S. , Chen Swaine L. , Hultgren Scott J.
Published: February 2010

Urinary tract infections (UTIs), the majority of which are caused by uropathogenicEscherichia coli(UPEC), afflict nearly 60% of women within their lifetimes. Studies in mice and humans have revealed that UPEC strains undergo a complex pathogenesis cycle that involves both the formation of intracellular bacterial communities (IBC) and the colonization of extracellular niches. Despite the commonality of the UPEC pathogenesis cycle, no specific urovirulence genetic profile has been determined; this is likely due to the fluid nature of the UPEC genome as the result of horizontal gene transfer and numerous genes of unknown function. UTI89 has a large extrachromosomal element termed pUTI89 with many characteristics of UPEC pathogenicity islands and that likely arose due to horizontal gene transfer. The pUTI89 plasmid has characteristics of both F plasmids and other known virulence plasmids. We sought to determine whether pUTI89 is important for virulence. Bothin vitroandin vivoassays were used to examine the function of pUTI89 using plasmid-cured UTI89. No differences were observed between UTI89 and plasmid-cured UTI89 based on growth, type 1 pilus expression, or biofilm formation. However, in a mouse model of UTI, a significant decrease in bacterial invasion, CFU and IBC formation of the pUTI89-cured strain was observed at early time points postinfection compared to the wild type. Through directed deletions of specific operons on pUTI89, thecjroperon was partially implicated in this observed defect. Our findings implicate pUTI89 in the early aspects of infection.
Local Host Response to Chlamydial Urethral Infection in Male Guinea Pigs
Authors: Wang Yin , Nagarajan Uma , Hennings Leah , Bowlin Anne K. , Rank Roger G.
Published: February 2010

Very little is known about the host response to chlamydial genital infection in the male, particularly about the nature of the local response in the urethra. In this study, the pathological and immunologic responses to urethral infection of the male guinea pig with
Chlamydia caviae
(
Chlamydophila caviae
) were characterized both during a primary infection and following a challenge infection. A dose-response experiment found that the 50% infectious dose for male urethral infection was 78 inclusion-forming units. The histopathologic response was similar to that of the female, with an initial acute inflammatory response followed by a chronic inflammatory response and plasma cell infiltration. Production of IgG and IgA antibodies in local urethral secretions developed following infection, and levels of both increased in a typical anamnestic response following a challenge infection. CD4 and CD8 T cells, as well as B cells, were observed in the local site by flow cytometry, with a slightly increased number of CD8 cells. Following challenge infection, the dominant anamnestic response was solely in the B-cell compartment, with only a minimal number of T cells. The T-cell response was clearly a Th1 response, as judged by increased levels of gamma interferon (IFN-γ), interleukin-12 p40 (IL-12p40), and IL-2. The proinflammatory cytokines and chemokines IL-8, IL-1β, tumor necrosis factor alpha (TNF-α), CCL2 (monocyte chemoattractant protein 1 [MCP-1]), and CCL5 (RANTES) were elicited in the urethra following primary infection, but only CCL5 showed increased levels upon challenge. This study represents the first comprehensive analysis of the local immune response in the male urethra to a chlamydial genital infection.
The diffuse double layer in ionic liquids
Authors: Fawcett W. Ronald , Ryan Peter J.
Published: February 2010

The equations used to describe the diffuse double layer in the Eigen–Wicke model of ionic liquids are presented. They are then used to estimate the potential drop across the diffuse layer and its differential capacity for two representative systems which contain monovalent ions of equal diameter. The first one is molten RbCl at 750 °C. The second system is a room temperature ionic liquid with typical parameters to describe its properties. The results of the calculations are compared with the available experimental data. It is concluded that the Eigen–Wicke model does not consider the change in local potential experienced by a given ion in the ionic liquid. The need for Monte Carlo data for the diffuse double layer in molten salt systems is emphasized.
Protofilament as a structural element of flagella of haloalkalophilic archaebacteria
Authors: Fedorov Oleg V. , Pyatibratov Michael G. , Kostyukova Alla S. , Osina Natalja K. , Tarasov Valery Yu.
Published: February 2010

Flagella of the haloalkalophilic archaebacterium Natronobacterium magadii were purified and characterized. The diameter of the flagella was 10 nm. It was shown that the flagella consist of four major proteins with molecular weights of 105 000, 60 000, 59 000, and 45 000. With decreasing NaCl concentration, the flagella dissociated into protofilaments. The structure of dissociated flagella and individual flagellins was studied by limited proteolysis. It was found that proteolytic cleavage of flagellins in dissociated flagella into high molecular weight fragments (about 40 000) did not lead to protofilament degradation. It was shown that the most stable fragment is formed from the 60 000 molecular weight flagellin. Cleavage of this fragment led to complete disappearance of protofilaments. On the basis of the data obtained, possible principles of archaebacterial flagellar construction are discussed.Key words: flagellin, archaebacteria, protofilaments, Natronobacterium magadii.
The absence of psychrophilic Thiobacillus ferrooxidans and acidophilic heterotrophic bacteria in cold, tailings effluents from a uranium mine
Authors: Berthelot Deborah , Leduc L. G. , Ferroni G. D.
Published: February 2010

Iron-oxidizing autotrophs (mainly Thiobacillus ferrooxidans) and acidophilic heterotrophs were recovered and quantified at an incubation temperature of 18 °C, in four tailings-effluent samples obtained from the environment of a uranium mine in Ontario, Canada. The samples were collected during winter when the temperatures of the effluents were in the range 0.5–5.0 °C. Iron-oxidizing autotrophs were recovered in the four samples and their numbers ranged from 3 ± 2 to 185 ± 18 colony-forming units/mL; acidophilic heterotrophs were recovered in three of the four samples and their numbers ranged from 13 ± 3 to 2517 ± 859 colony-forming units/mL. Forty-six of the iron-oxidizer isolates and 63 of the acidophilic heterotrophic isolates were examined for their ability to grow at temperatures of 4, 18, 21, and 37 °C. None of the isolates was psychrophilic, although 96% of the iron oxidizers and 54% of the acidophilic heterotrophs were psychrotrophs; less that 5% of the isolates for both nutritional types were capable of growth at 37 °C. In addition, the isolates were categorized as 'broader temperature range psychrotrophs,' 'narrower temperature range psychrotrophs,' 'intermediates,' or mesophiles, and the narrower temperature range psychrotrophs were found to be numerically predominant.Key words: psychrotrophs, psychrophiles, Thiobacillus ferrooxidans, acidophilic heterotrophic bacteria.
Alterations in total bacteria, iodonitrophenyltetrazolium (INT)-positive bacteria, and heterotrophic plate counts of bottled mineral water
Authors: Ferreira Ana-Cristina , Morais Paula Vasconcellos , Costa Milton S. da
Published: February 2010

The quantitative variation in the microflora in bottled mineral waters stored in polyvinyl chloride bottles was studied immediately after bottling and after 7 days storage, over 9 months. The microflora was determined by combining total counts using ethidium bromide, and the number of iodonitrophenyltetrazolium (INT)-positive cells determined with heterotrophic plate counts on R2A for prolonged incubation periods at 22 and 37 °C. Immediately after bottling of mineral water A, the total counts with ethidium bromide varied between 70.0 and 495.0 bacteria/mL. The number of INT-positive bacteria varied between 14.0 and 40.0% counts. The heterotrophic plate counts were higher at 22 °C than at 37 °C and varied between 1.1 and 13.2% and between 0.7 and 9.3% of the total counts, respectively. After 7 days storage, total bacterial counts increased by 1000 times, indicating multiplication of bacteria that were present in low numbers immediately after bottling. The percentage of INT-positive bacteria remained between 2.6 and 39.0% of the total counts but the number of culturable bacteria was higher than the number of INT-positive cells. Similar counts were found in other mineral waters (B, C, and D), but the numbers of INT-positive cells in two of them (B and D) were higher than the heterotrophic plate counts at 22 °C. These results show the presence in the aquifer and (or) bottling system of a flora that is neither INT-positive nor culturable and reinforce previous results that still mineral waters have a large bacterial population after storage.Key words: mineral water, bacterial populations, ethidium bromide, INT, HPC.
Adherence of Candida albicans germ tubes to murine tissues in an ex vivo assay
Authors: López-Ribot Jose Luis , Vespa Maria Novella , Chaffin W. LaJean
Published: February 2010

Adhesion of Candida albicans germ tubes to murine tissues was examined. An ex vivo assay previously employed to examine adhesion of yeast cells of C. albicans was adapted for use with germ tubes. Binding of germ tubes to kidney, liver, spleen, and lymph node tissues was found to occur throughout the tissue section, with little tissue morphologic specificity. In general, more organisms adhered to spleen and lymph node tissues than to kidney and liver tissues. Observation of adhesion with scanning electron microscopy showed three germ tube – tissue interactions described as loose, tight, or embedded.Key words: Candida, germ tubes, adhesion, ex vivo.
Electroporation and its effect on the psychrotrophic bacterium Bacillus psychrophilus
Authors: Whyte Lyle G. , Inniss William E.
Published: February 2010

The psychrotrophic bacterium Bacillus psychrophilus was successfully transformed by electroporation with the cloning vector pC194 and the expression vector pPL708. Optimal electroporation parameters such as field strength, pulse length, and electroporation medium, as well as the influence of the growth phase of the bacterial cells, were determined. Maximum transformation efficiencies of 104 transformants/μg plasmid DNA were achieved with late logarithmic – early stationary phase cells grown in medium supplemented with 33.3 mM glycine and electroporated at high field strengths of 18–20 kV∙cm−1. By means of this procedure, a kanamycin-resistant gene was directly cloned into pPL708 and the recombinant plasmid electrotransformed into B. psychrophilus. In addition, the effect of electroporation on protein synthesis was analyzed by 2-dimensional polyacrylamide gel electrophoresis and computing scanning laser densitometry. At least eight proteins were induced by electroporation field strengths of 18 or 25 kV during the first 2 h immediately following electroporation. Conversely, the synthesis of at least five proteins was repressed by electroporation.Key words: electroporation, psychrotrophic bacterium.