bluuebunny/crossref_metadata_2025 huggingface dataset:
Reliable assessment of forearm photoageing by high‐frequency ultrasound: a cross‐sectional study
Authors: Caetano L. de V. N. , Soares J. L. M. , Bagatin E. , Miot H. A.
Published: September 2015

Synopsis

Objective

High‐frequency ultrasound is a non‐invasive tool used in skin ageing research to assess dermis thickness and echogenicity. This study evaluated the reliability of a range of high‐frequency ultrasound parameters and tested their correlation with age and a validated clinical scale for the assessment of forearm skin photoageing; the difference between two body sites according to environmental exposition patterns was also investigated.

Methods

Twenty‐three volunteers aged 28–82 years were divided into three groups according to forearm photoageing degree. A 20 MHz ultrasound unit was used to obtain cross‐sectional images of the skin by two trained investigators on two different sites: the dorsal forearm (chronically photoexposed skin) and the proximal medial arm (non‐photoexposed skin). Several echogenicity parameters were studied for each skin compartment: total dermis, upper dermis and lower dermis, and the ratio between upper and lower dermis.

Results

The intraclass correlation coefficient (for complete agreement) between investigators was higher for upper and total dermis echogenicity measures compared with the lower dermis. At the non‐photoexposed site, the upper and lower dermis parameter ratio was better correlated with age. At the photoexposed area, total dermis parameters demonstrated higher correlations with clinical score.

Conclusion

The authors discuss the choice of parameters for forearm photoageing assessment using high‐frequency ultrasound.
Rheological properties, ginsenosides contents, sensory evaluations of Korean red ginseng extracts
Authors: Chang Yoon Hyuk , Kwak Han Sub , Yoo Byoungseung , Lee Youngseung
Published: September 2015

Red ginseng extracts have gained popularity due to their positive impact on health. The objective of this study was to identify the rheological properties, ginsenosides contents and sensory characteristics of six different commercial red ginseng extracts in Korea. Ginseng extracts demonstrated different shear‐thinning flow behaviour (n = 0.59–0.78) with a consistency index (K = 46.77–105.78 Pa sn), which meant they had different viscosity. The amount of ginsenosides and total solid contents were highly correlated (r = 0.954). Six significant attributes (ginseng, angelica, bitterness, ginger flavour and astringency) were identified and can be used as sensory parameters for the determination of the product quality. Hedonic levels of the product were assessed by a group of ninety‐five consumers in their 20 s (n = 32) and over 30 (n = 63). Cluster analysis revealed that consumer acceptance was divided into two major clusters for those that preferred strong (n = 43) and weak (n = 42) ginseng characteristics in the extract.
Need for a roadmap for development of a coordinated national registry programme
Authors: Wilkins S. , Best R. L. , Evans S. M.
Published: September 2015

Clinical quality registries are an overlooked and under‐funded arm of clinical research in Australia. Registries are databases for patients with a particular disease, or who undergo a procedure, or use a health resource. Registries, where properly funded and universally adopted, have provided substantial benefits to the quality of healthcare and, in some cases, have had demonstrable effect in reducing costs. There is a lack of a coordinated programme for both funding and development of registries in Australia. A coordinated effort is required to address key gaps in registry coverage and ensure registries comply with appropriate technical and operating principles, and target areas where registries can add value to the health system. This will ensure that Australia is competitive with its international peers in this dynamic environment.
Attitudes of midwives and maternal child health nurses towards suicide: A cross‐sectional study
Authors: Lau Rosalind , McCauley Kay , Barnfield Jakqui , Moss Cheryle , Cross Wendy
Published: September 2015

Perinatal women are at risk of depression and/or suicidality. Suicide is the highest cause of indirect maternal deaths in the perinatal period. Midwives and maternal child health nurses (MCHN), as key clinicians, need to be able to detect these mental health issues. Little is known about these clinicians' attitudes to suicide. In this paper, we report on the results of a cross‐sectional study of midwives' and MCHN attitudes to suicide. A convenience sample of midwives (n = 95) and MCHN (n = 86) from south–eastern Victoria, Australia, was recruited into the study. Participants completed the Attitudes to Suicide Prevention Scale. The results showed that MCHN have more positive attitudes towards suicide prevention than midwives, and younger participants have more positive attitudes to suicide prevention compared to older participants. Midwives and MCHN could benefit from continuing professional education to build their knowledge and skills in assessing suicide risk for childbearing women and their families, increasing positive attitudes, improving detection, and mental health referrals.
Animal‐inflicted open wounds in rural Turkey: lessons learned and a proposed treatment algorithm for uncertain scenarios
Authors: Sezgin Billur , Ljohiy Mbaraka , Akgol Gur Sultan Tuna
Published: September 2015

Uncertainty in the management of animal‐inflicted injuries, especially in rural settings, usually results in a general approach to leave all wounds to heal with secondary intention, which can lead to unsightly scarring and functional loss. This study focusus on different circumstances dealt with by plastic surgeons in a rural setting in Turkey and aims to configure what the general approach should be through an analysis of a wide spectrum of patients.

Between June 2013 and December 2014, 205 patients who presented to the emergency department for animal‐inflicted injuries were retrospectively analysed. Patients who consulted for plastic surgery were included in the analysis to determine which wounds require further attention. Patients with past animal‐inflicted injuries who presented to the outpatient plastic surgery clinic with concerns such as non‐healing open wounds or cosmetic or functional impairment were also evaluated.

Statistical analysis demostrated a significantly lower rate of infection encountered in animal‐inflicted open wounds (AIOWs) of patients who consulted for plastic surgery from the emergency department than those who presented to the outpatient clinic (P < 0·05).

The main concern in the management of animal‐inflicted wounds is their potential for infection, but this does not mean that every wound will be infected. The most important factor is being able to distinguish wounds that have a higher potential for infection and to select the type of wound management accordingly. An algorithm has been proposed as a guidance for the management of AIOWs, which covers the approach towards both domestic and stray animal‐inflicted injuries.
Population structure within an alpine archipelago: strong signature of past climate change in the New Zealand rock wren (Xenicus gilviventris)
Authors: Weston K. A. , Robertson B. C.
Published: September 2015

Naturally subdivided populations such as those occupying high‐altitude habitat patches of the ‘alpine archipelago’ can provide significant insight into past biogeographical change and serve as useful models for predicting future responses to anthropogenic climate change. Among New Zealand's alpine taxa, phylogenetic studies support two major radiations: the first correlating with geological forces (Pliocene uplift) and the second with climatic processes (Pleistocene glaciations). The rock wren (Xenicus gilviventris) is a threatened alpine passerine belonging to the endemic New Zealand wren family (Acanthisittidae). Rock wren constitute a widespread, naturally fragmented population, occurring in patches of suitable habitat over c. 900 m in altitude throughout the length of the South Island, New Zealand. We investigated the relative role of historical geological versus climatic processes in shaping the genetic structure of rock wren (N = 134) throughout their range. Using microsatellites combined with nuclear and mtDNA sequence data, we identify a deep north–south divergence in rock wren (3.7 ± 0.5% at cytochrome b) consistent with the glacial refugia hypothesis whereby populations were restricted in isolated refugia during the Pleistocene c. 2 Ma. This is the first study of an alpine vertebrate to test and provide strong evidence for the glacial refugia hypothesis as an explanation for the low endemicity central zone known as the biotic ‘gap’ in the South Island of New Zealand.
Back to the Future?: Emergent Visions for Object‐Based Teaching in and beyond the Classroom
Authors: Adams Kathleen M.
Published: September 2015

This article introduces a special issue ofMuseum Anthropologydevoted to innovative strategies for teaching with objects. Although a century ago anthropology, museums, and objects were intimately entwined, trends in many museology and anthropology courses have drifted toward focusing on ideas and people rather than objects. The contributors to this special issue have cultivated new pedagogical approaches that complement or realign literature‐focused classroom canons that can distance students from the very objects under study. In keeping with recent theoretical approaches to objects that highlight the sensory dimensions of material culture, many of the articles in this special issue examine the challenges and potential rewards when educators foster physical engagement with objects in and beyond the classroom. Taken together, the articles also underscore how object‐based teaching can yield new theoretical and practical insights, enhance the social relevance of classroom activities, and facilitate meaningful benefits for local communities. [material culture, praxis, pedagogy, embodied practices, object‐based teaching]
Two cases of rheumatoid meningitis
Authors: Magaki Shino , Chang Edward , Hammond Robert R. , Yang Isaac , Mackenzie Ian R. A. , Chou Benedict T. , Choi Soo I. , Jen Joanna C. , Pope Whitney B. , Bell David A. , Vinters Harry V.
Published: September 2015

Central nervous system (CNS) involvement by rheumatoid arthritis (RA) in the form of rheumatoid meningitis (RM) is rare and most commonly occurs in the setting of longstanding severe RA. Due to a wide range of clinical presentations and nonspecific laboratory findings, it presents a diagnostic challenge often requiring brain biopsy. Only a few histopathologically confirmed cases have been described in the literature. Our aim is to describe two cases of RM and review the literature. The first case is of a previously healthy 37‐year‐old man who presented with severe headaches and focal neurologic deficits. Magnetic resonance imaging demonstrated abnormal leptomeningeal enhancement in the left frontal and parietal sulci. The second case is of a 62‐year‐old woman with a history of mild chronic joint pain who presented with confusion, personality changes and seizures. Both patients ultimately underwent brain biopsy which demonstrated RM on pathologic examination. Administration of corticosteroids resulted in significant clinical improvement in both cases. To our knowledge, our unusual case of RM in the young man is the fifth reported case of rheumatoid meningitis in a patient with no prior history of RA. Such an atypical presentation makes diagnosis even more difficult and highlights the need for awareness of this entity in the diagnostic consideration of a patient presenting with unexplained neurologic symptoms. Our literature review underscores the clinical and pathologic heterogeneity of CNS involvement in RA.
Flavins secreted by roots of iron‐deficient Beta vulgaris enable mining of ferric oxide via reductive mechanisms
Authors: Sisó‐Terraza Patricia , Rios Juan J. , Abadía Javier , Abadía Anunciación , Álvarez‐Fernández Ana
Published: September 2015

Iron (Fe) is abundant in soils but generally poorly soluble. Plants, with the exception of Graminaceae, take up Fe using an Fe(III)‐chelate reductase coupled to an Fe(II) transporter. Whether or not nongraminaceous species can convert scarcely soluble Fe(III) forms into soluble Fe forms has deserved little attention so far. We have used Beta vulgaris, one among the many species whose roots secrete flavins upon Fe deficiency, to study whether or not flavins are involved in Fe acquisition.

Flavins secreted by Fe‐deficient plants were removed from the nutrient solution, and plants were compared with Fe‐sufficient plants and Fe‐deficient plants without flavin removal. Solubilization of a scarcely soluble Fe(III)‐oxide was assessed in the presence or absence of flavins, NADH (nicotinamide adenine dinucleotide, reduced form) or plant roots, and an Fe(II) trapping agent.

The removal of flavins from the nutrient solution aggravated the Fe deficiency‐induced leaf chlorosis. Flavins were able to dissolve an Fe(III)‐oxide in the presence of NADH. The addition of extracellular flavins enabled roots of Fe‐deficient plants to reductively dissolve an Fe(III)‐oxide.

We concluded that root‐secretion of flavins improves Fe nutrition in B. vulgaris. Flavins allow B. vulgaris roots to mine Fe from Fe(III)‐oxides via reductive mechanisms.
Inhibition of LtxA toxicity by blocking cholesterol binding with peptides
Authors: Brown A.C. , Koufos E. , Balashova N.V. , Boesze‐Battaglia K. , Lally E.T.
Published: September 2015

The leukotoxin (LtxA) produced by Aggregatibacter actinomycetemcomitans kills host immune cells, allowing the bacterium to establish an ecological niche in the upper aerodigestive tract of its human host. The interaction of LtxA with human immune cells is both complex and multifaceted, involving membrane lipids as well as cell‐surface proteins. In the initial encounter with the host cell, LtxA associates with lymphocyte function‐associated antigen‐1, a cell surface adhesion glycoprotein. However, we have also demonstrated that the toxin associates strongly with the plasma membrane lipids, specifically cholesterol. This association with cholesterol is regulated by a cholesterol recognition amino acid consensus (CRAC) motif, with a sequence of 334LEEYSKR340, in the N‐terminal region of the toxin. Here, we have demonstrated that removal of cholesterol from the plasma membrane or mutation of the LtxA CRAC motif inhibits the activity of the toxin in THP‐1 cells. To inhibit LtxA activity, we designed a short peptide corresponding to the CRAC336 motif of LtxA (CRAC336WT). This peptide binds to cholesterol and thereby inhibits the toxicity of LtxA in THP‐1 cells. Previously, we showed that this peptide inhibits LtxA toxicity against Jn.9 (Jurkat) cells, indicating that peptides derived from the cholesterol‐binding site of LtxA may have a potential clinical applicability in controlling infections of repeats‐in‐toxin‐producing organisms.
Multidimensional anatomy of ‘modern type depression’ in Japan: A proposal for a different diagnostic approach to depression beyond the DSM‐5
Authors: Kato Takahiro A. , Hashimoto Ryota , Hayakawa Kohei , Kubo Hiroaki , Watabe Motoki , Teo Alan R. , Kanba Shigenobu
Published: September 2015

Japan's prototype of depression was traditionally a melancholic depression based on the premorbid personality known as shūchaku‐kishitsu proposed by Mitsuzo Shimoda in the 1930s. However, since around 2000, a novel form of depression has emerged among Japanese youth. Called ‘modern type depression (MTD)’ by the mass media, the term has quickly gained popularity among the general public, though it has not been regarded as an official medical term. Likewise, lack of consensus guidelines for its diagnosis and treatment, and a dearth of scientific literature on MTD has led to confusion when dealing with it in clinical practice in Japan. In this review article, we summarize and discuss the present situation and issues regarding MTD by focusing on historical, diagnostic, psychosocial, and cultural perspectives. We also draw on international perspectives that begin to suggest that MTD is a phenomenon that may exist not only in Japan but also in many other countries with different sociocultural and historical backgrounds. It is therefore of interest to establish whether MTD is a culture‐specific phenomenon in Japan or a syndrome that can be classified using international diagnostic criteria as contained in the ICD or the DSM. We propose a novel diagnostic approach for depression that addresses MTD in order to combat the current confusion about depression under the present diagnostic systems.
Development of ensemble pedotransfer functions for cation exchange capacity of soils ofQingdao inChina
Authors: Liao K. , Xu S. , Zhu Q.
Published: September 2015

Soil cation exchange capacity (CEC), which is considered to be an indicator of buffering capacity, is an important soil attribute that influences soil fertility but is costly, time‐consuming and labour‐intensive to measure. Pedotransfer functions (PTFs) have routinely been used to predict soilCECfrom easily measured soil properties, such as soilpH, texture and organic matter content. However, uncertainty in which one to select can be substantial as differentPTFs do not necessarily produce the same result. In this study, a total of 100 soil samples were collected from surface horizons (0–20 cm) in different regions of Qingdao City, China. Three ensemblePTFs (ePTFs), including simple ensemble mean (SEM), individually bias‐removed ensemble mean (IBREM) and collective bias‐removed ensemble mean (CBREM), were developed to reduce the uncertainty inCECprediction based on 12 published regression‐basedPTFs. In addition, a localPTF(LPTF) forCECwas also developed using multiple stepwise regression and basic soil properties. The performances of the threeePTFs were compared with those of the publishedPTFs andLPTF. Results show that the differences between the performances of the publishedPTFs were substantial. When the systematic bias of each publishedPTFwas removed separately, the prediction capability of thePTFs was increased. The performance ofLPTFwas significantly better than that ofSEM, but slightly worse thanIBREM. It is noted thatCBREMhad higher accuracy than all of the other methods. Overall,CBREMis a promising approach for estimating soilCECin the study area.
An index theorem for end-periodic operators
Authors: Mrowka Tomasz , Ruberman Daniel , Saveliev Nikolai
Published: September 2015

We extend the Atiyah, Patodi, and Singer index theorem for first-order differential operators from the context of manifolds with cylindrical ends to manifolds with periodic ends. This theorem provides a natural complement to Taubes’ Fredholm theory for general end-periodic operators. Our index theorem is expressed in terms of a new periodic eta-invariant that equals the Atiyah–Patodi–Singer eta-invariant in the cylindrical setting. We apply this periodic eta-invariant to the study of moduli spaces of Riemannian metrics of positive scalar curvature.
Perivascular tissue inhibits rho‐kinase‐dependent smooth muscle Ca2+ sensitivity and endothelium‐dependent H2S signalling in rat coronary arteries
Authors: Aalbaek Filip , Bonde Lisbeth , Kim Sukhan , Boedtkjer Ebbe
Published: September 2015

Key points

Local regulation of vascular resistance adjusts coronary blood flow to metabolic demand, although the mechanisms involved are not comprehensively understood

We show that heart tissue surrounding rat coronary arteries releases diffusible factors that regulate vasoconstriction and relaxation

Perivascular tissue reduces rho‐kinase‐dependent smooth muscle Ca2+ sensitivity and constriction of coronary arteries to serotonin, the thromboxane analogue U46619 and the α1‐adrenergic agonist phenylephrine

Endothelium‐dependent relaxation of coronary arteries in response to cholinergic stimulation is inhibited by perivascular tissue as a result of reduced endothelial Ca2+ responses and attenuated H2S‐dependent signalling

These results establish cellular mechanisms by which perivascular heart tissue can modify local vascular tone and coronary blood flow

Interactions between perivascular tissue (PVT) and the vascular wall modify artery tone and contribute to local blood flow regulation. Using isometric myography, fluorescence microscopy, membrane potential recordings and phosphospecific immunoblotting, we investigated the cellular mechanisms by which PVT affects constriction and relaxation of rat coronary septal arteries. PVT inhibited vasoconstriction to thromboxane, serotonin and α1‐adrenergic stimulation but not to depolarization with elevated extracellular [K+]. When PVT was wrapped around isolated arteries or placed at the bottom of the myograph chamber, a smaller yet significant inhibition of vasoconstriction was observed. Resting membrane potential, depolarization to serotonin or thromboxane stimulation, and resting and serotonin‐stimulated vascular smooth muscle [Ca2+]‐levels were unaffected by PVT. Serotonin‐induced vasoconstriction was almost abolished by rho‐kinase inhibitor Y‐27632 and modestly reduced by protein kinase C inhibitor bisindolylmaleimide X. PVT reduced phosphorylation of myosin phosphatase targeting subunit (MYPT) at Thr850 by ∼40% in serotonin‐stimulated arteries but had no effect on MYPT‐phosphorylation in arteries depolarized with elevated extracellular [K+]. The net anti‐contractile effect of PVT was accentuated after endothelial denudation. PVT also impaired vasorelaxation and endothelial Ca2+ responses to cholinergic stimulation. Methacholine‐induced vasorelaxation was mediated by NO and H2S, and particularly the H2S‐dependent (dl‐propargylglycine‐ and XE991‐sensitive) component was attenuated by PVT. Vasorelaxation to NO‐ and H2S‐donors was maintained in arteries with PVT. In conclusion, cardiomyocyte‐rich PVT surrounding coronary arteries releases diffusible factors that reduce rho‐kinase‐dependent smooth muscle Ca2+ sensitivity and endothelial Ca2+ responses. These mechanisms inhibit agonist‐induced vasoconstriction and endothelium‐dependent vasorelaxation and suggest new signalling pathways for metabolic regulation of blood flow.
Field Comparison of IEC 61400-11 Wind Turbines — Part 11: Acoustic Noise Measurement Techniques, Edition 3.0 and Edition 2.1
Authors: Jozwiak Rob , Munro Allan , Halstead Duncan , Denison Addie
Published: September 2015

Following the release of Edition 3.0 (2012) of IEC 61400-11 Wind Turbines – Part 11: Acoustic noise measurement techniques, there has been a lot of interest as to how analysis results differ from methods stipulated in Edition 2.1 (2006).

This paper provides a detailed review of the differences between Edition 3.0 and Edition 2.1. An analysis is provided on differences in evaluation of the apparent sound power level and tonal audibility between both versions of the measurement standard.
Technical Note: Synchrotron‐based high‐energy x‐ray phase sensitive microtomography for biomedical research
Authors: Liu Huiqiang , Wu Xizeng , Xiao Tiqiao
Published: September 2015

Purpose:

Propagation‐based phase‐contrast CT (PPCT) utilizes highly sensitive phase‐contrast technology applied to x‐ray microtomography. Performing phase retrieval on the acquired angular projections can enhance image contrast and enable quantitative imaging. In this work, the authors demonstrate the validity and advantages of a novel technique for high‐resolution PPCT by using the generalized phase‐attenuation duality (PAD) method of phase retrieval.

Methods:

A high‐resolution angular projection data set of a fish head specimen was acquired with a monochromatic 60‐keV x‐ray beam. In one approach, the projection data were directly used for tomographic reconstruction. In two other approaches, the projection data were preprocessed by phase retrieval based on either the linearized PAD method or the generalized PAD method. The reconstructed images from all three approaches were then compared in terms of tissue contrast‐to‐noise ratio and spatial resolution.

Results:

The authors’ experimental results demonstrated the validity of the PPCT technique based on the generalized PAD‐based method. In addition, the results show that the authors’ technique is superior to the direct PPCT technique as well as the linearized PAD‐based PPCT technique in terms of their relative capabilities for tissue discrimination and characterization.

Conclusions:

This novel PPCT technique demonstrates great potential for biomedical imaging, especially for applications that require high spatial resolution and limited radiation exposure.
Correcting scan‐to‐scan response variability for a radiochromic film‐based reference dosimetry system
Authors: Lewis David , Devic Slobodan
Published: September 2015

Purpose:

In radiochromic film dosimetry systems, measurements are usually obtained from film images acquired on a CCD‐based flatbed scanner. The authors investigated factors affecting scan‐to‐scan response variability leading to increased dose measurement uncertainty.

Methods:

The authors used flatbed document scanners to repetitively scan EBT3 radiochromic films exposed to doses 0–1000 cGy, together with three neutral density filters and three blue optical filters. Scanning was performed under two conditions: scanner lid closed and scanner lid opened/closed between scans. The authors also placed a scanner in a cold room at 9 °C and later in a room at 22 °C and scanned EBT3 films to explore temperature effects. Finally, the authors investigated the effect of altering the distance between the film and the scanner's light source.

Results:

Using a measurement protocol to isolate the contribution of the CCD and electronic circuitry of the scanners, the authors found that the standard deviation of response measurements for the EBT3 film model was about 0.17% for one scanner and 0.09% for the second. When the lid of the first scanner was opened and closed between scans, the average scan‐to‐scan difference of responses increased from 0.12% to 0.27%. Increasing the sample temperature during scanning changed the RGB response values by about −0.17, −0.14, and −0.05%/°C, respectively. Reducing the film‐to‐light source distance increased the RBG response values about 1.1, 1.3, and 1.4%/mm, respectively. The authors observed that films and film samples were often not flat with some areas up to 8 mm away from the scanner's glass window.

Conclusions:

In the absence of measures to deal with the response irregularities, each factor the authors investigated could lead to dose uncertainty >2%. Those factors related to the film‐to‐light source distance could be particularly impactful since the authors observed many instances where the curl of film samples had the potential to cause dose uncertainty in excess of 5%. Two expedients will eliminate the uncertainties: a transparent sheet (preferably glass) placed over the scanned film keeps the film‐to‐light source distance constant, and an EBT3 reference film included in all scans provides correction factors for measured response values.
Tumor cell dispersal: A little movement goes a long way
Authors: Lalazar Gadi
Published: September 2015

In a spatial model, short-range dispersal of tumor cells has a major impact on limiting tumor genetic heterogeneity, tumor growth rate, and resistance to chemotherapy.
Eagle Effect in Nonreplicating Persister Mycobacteria
Authors: Wu Mu-Lu , Tan Jasmie , Dick Thomas
Published: September 2015

We determined the microbicidal activities of antibacterials against nonreplicating
Mycobacterium smegmatis
grown in a starvation-based Loebel model for persistence. Whereas most drugs lost their activity, fluoroquinolones retained lethal potency. Dose-response characterizations showed a paradoxical more-drug-kills-less Eagle effect. Pretreatment of cultures with chloramphenicol blocked the lethal action of the gyrase inhibitors. These results suggest that fluoroquinolones at low concentrations trigger a protein synthesis-dependent cell death pathway and shut off this suicide pathway at elevated concentrations.
A Novel Pyridazinone Derivative Inhibits Hepatitis B Virus Replication by Inducing Genome-Free Capsid Formation
Authors: Wang Ya-Juan , Lu Dong , Xu Yi-Bin , Xing Wei-Qiang , Tong Xian-Kun , Wang Gui-Feng , Feng Chun-Lan , He Pei-Lan , Yang Li , Tang Wei , Hu You-Hong , Zuo Jian-Ping
Published: September 2015

Here we first identified a novel pyridazinone derivative, compound 3711, as a nonnucleosidic hepatitis B virus (HBV) inhibitor in a cell model system. 3711 decreased extracellular HBV DNA levels by 50% (50% inhibitory concentration [IC
50
]) at 1.5 ± 0.2 μM and intracellular DNA levels at 1.9 ± 0.1 μM, which demonstrated antiviral activity at levels far below those associated with toxicity. Both the 3TC/ETV dually resistant L180M/M204I mutant and the adefovir (ADV)-resistant A181T/N236T mutant were as susceptible to 3711 as wild-type HBV. 3711 treatment induced the formation of genome-free capsids, a portion of which migrated faster on 1.8% native agarose gel. The induced genome-free capsids sedimented more slowly in isopycnic CsCl gradient centrifugation without significant morphological changes. 3711 treatment decreased levels of HBV DNA contained in both secreted enveloped virion and naked virus particles in supernatant. 3711 could interfere with capsid formation of the core protein (Cp) assembly domain. A Cp V124W mutant, which strengthens capsid interdimer interactions, recapitulated the effect of 3711 on capsid assembly. Pyridazinone derivative 3711, a novel chemical entity and HBV inhibitor, may provide a new opportunity to combat chronic HBV infection.
A Clonal Complex 12 Methicillin-Resistant Staphylococcus aureus Strain, West Australian MRSA-59, Harbors a Novel Pseudo-SCC
mec
Element
Authors: Monecke Stefan , Coombs Geoffrey W. , Pearson Julie , Hotzel Helmut , Slickers Peter , Ehricht Ralf
Published: September 2015

A West Australian methicillin-resistant
Staphylococcus aureus
strain (WA MRSA-59) was characterized by microarray and sequencing. Its pseudo-staphylococcal cassette chromosome
mec
(SCC
mec
) element comprised
dcs
,
Q9XB68
-
dcs
,
mvaS
-SCC,
Q5HJW6
,
dru
,
ugpQ
,
ydeM
,
mecA-mecR-mecI
, txbi
mecI
,
tnp
IS
431
,
copA2-mco
(copper resistance),
ydhK
,
arsC-arsB-arsR
(arsenic resistance), open reading frame PT43, and
per-2
. Recombinase genes,
xylR
(
mecR2
), and PSM-
mec
(phenol-soluble modulin) were absent. We suggest that
mec
complex A should be split into two subtypes. One harbors PSM-
mec
and
xylR
(
mecR2
). It is found in SCC
mec
types II, III, and VIII. The second subtype, described herein, is present in WA MRSA-59 and some coagulase-negative staphylococci.
Physiological Roles and Adverse Effects of the Two Cystine Importers of Escherichia coli
Authors: Chonoles Imlay Karin R. , Korshunov Sergey , Imlay James A.
Published: September 2015

When cystine is added to
Escherichia coli
, the bacterium becomes remarkably sensitive to hydrogen peroxide. This effect is due to enlarged intracellular pools of cysteine, which can drive Fenton chemistry. Genetic analysis linked the sensitivity to YdjN, a secondary transporter that along with the FliY-YecSC ABC system is responsible for cystine uptake. FliY-YecSC has a nanomolar

K
m

and is essential for import of trace cystine, whereas YdjN has a micromolar

K
m

and is the predominant importer when cystine is more abundant. Oddly, both systems are strongly induced by the CysB response to sulfur scarcity. The FliY-YecSC system can import a variety of biomolecules, including diaminopimelate; it is therefore vulnerable to competitive inhibition, presumably warranting YdjN induction under low-sulfur conditions. But the consequence is that if micromolar cystine then becomes available, the abundant YdjN massively overimports it, at >30 times the total sulfur demand of the cell. The imported cystine is rapidly reduced to cysteine in a glutathione-dependent process. This action avoids the hazard of disulfide stress, but it precludes feedback inhibition of YdjN by cystine. We conjecture that YdjN possesses no cysteine allosteric site because the isostructural amino acid serine might inappropriately bind in its place. Instead, the cell partially resolves the overaccumulation of cysteine by immediately excreting it, completing a futile import/reduction/export cycle that consumes a large amount of cellular energy. These unique, wasteful, and dangerous features of cystine metabolism are reproduced by other bacteria. We propose to rename
ydjN
as
tcyP
and
fliY-yecSC
as
tcyJLN
.

IMPORTANCE
In general, intracellular metabolite pools are kept at steady, nontoxic levels by a sophisticated combination of transcriptional and allosteric controls. Surprisingly, in
E. coli
allosteric control is utterly absent from the primary importer of cystine. This flaw allows massive overimport of cystine, which causes acute vulnerability to oxidative stress and is remedied only by wasteful cysteine efflux. The lack of import control may be rationalized by the unusual properties of cysteine itself. This phenomenon justifies the existence of countervailing cysteine export systems, whose purpose is otherwise hard to understand. It also highlights an unexpected link between sulfur metabolism and oxidative damage. Although this investigation focused upon
E. coli
, experiments confirmed that similar phenomena occur in other species.
Characterization of 3′-Phosphate RNA Ligase Paralogs RtcB1, RtcB2, and RtcB3 from Myxococcus xanthus Highlights DNA and RNA 5′-Phosphate Capping Activity of RtcB3
Authors: Maughan William P. , Shuman Stewart
Published: September 2015

Escherichia coli
RtcB exemplifies a family of GTP-dependent RNA repair/splicing enzymes that join 3′-PO
4
ends to 5′-OH ends via stable RtcB-(histidinyl-
N
)-GMP and transient RNA
3′
pp
5′
G intermediates.
E. coli
RtcB also transfers GMP to a DNA 3′-PO
4
end to form a stable “capped” product, DNA
3′
pp
5′
G. RtcB homologs are found in a multitude of bacterial proteomes, and many bacteria have genes encoding two or more RtcB paralogs; an extreme example is
Myxococcus xanthus
, which has six RtcBs. In this study, we purified, characterized, and compared the biochemical activities of three
M. xanthus
RtcB paralogs. We found that
M. xanthus
RtcB1 resembles
E. coli
RtcB in its ability to perform intra- and intermolecular sealing of a
HO
RNAp substrate and capping of a DNA 3′-PO
4
end.
M. xanthus
RtcB2 can splice
HO
RNAp but has 5-fold-lower RNA ligase specific activity than RtcB1. In contrast,
M. xanthus
RtcB3 is distinctively feeble at ligating the
HO
RNAp substrate, although it readily caps a DNA 3′-PO
4
end. The novelty of
M. xanthus
RtcB3 is its capacity to cap DNA and RNA 5′-PO
4
ends to form GppDNA and GppRNA products, respectively. As such, RtcB3 joins a growing list of enzymes (including RNA 3′-phosphate cyclase RtcA and thermophilic ATP-dependent RNA ligases) that can cap either end of a polynucleotide substrate. GppDNA formed by RtcB3 can be decapped to pDNA by the DNA repair enzyme aprataxin.

IMPORTANCE
RtcB enzymes comprise a widely distributed family of RNA 3′-PO
4
ligases distinguished by their formation of 3′-GMP-capped RNAppG and/or DNAppG polynucleotides. The mechanism and biochemical repertoire of
E. coli
RtcB are well studied, but it is unclear whether its properties apply to the many bacteria that have genes encoding multiple RtcB paralogs. A comparison of the biochemical activities of three
M. xanthus
paralogs, RtcB1, RtcB2, and RtcB3, shows that not all RtcBs are created equal. The standout findings concern RtcB3, which is (i) inactive as an RNA 3′-PO
4
ligase but adept at capping a DNA 3′-PO
4
end and (ii) able to cap DNA and RNA 5′-PO
4
ends to form GppDNA and GppRNA, respectively. The GppDNA and GppRNA capping reactions are novel nucleic acid modifications.
Association with PAK2 Enables Functional Interactions of Lentiviral Nef Proteins with the Exocyst Complex
Authors: Imle Andrea , Abraham Libin , Tsopoulidis Nikolaos , Hoflack Bernard , Saksela Kalle , Fackler Oliver T.
Published: September 2015

Human immunodeficiency virus type 1 (HIV-1) Nef enhances virus replication and contributes to immune evasion
in vivo
, but the underlying molecular mechanisms remain incompletely defined. Nef interferes with host cell actin dynamics to restrict T lymphocyte responses to chemokine stimulation and T cell receptor engagement. This relies on the assembly of a labile multiprotein complex including the host kinase PAK2 that Nef usurps to phosphorylate and inactivate the actin-severing factor cofilin. Components of the exocyst complex (EXOC), an octameric protein complex involved in vesicular transport and actin remodeling, were recently reported to interact with Nef via the same molecular surface that mediates PAK2 association. Exploring the functional relevance of EXOC in Nef-PAK2 complex assembly/function, we found Nef-EXOC interactions to be specifically mediated by the PAK2 interface of Nef, to occur in infected human T lymphocytes, and to be conserved among lentiviral Nef proteins. In turn, EXOC was dispensable for direct downstream effector functions of Nef-associated PAK2. Surprisingly, PAK2 was essential for Nef-EXOC association, which required a functional Rac1/Cdc42 binding site but not the catalytic activity of PAK2. EXOC was dispensable for Nef functions in vesicular transport but critical for inhibition of actin remodeling and proximal signaling upon T cell receptor engagement. Thus, Nef exploits PAK2 in a stepwise mechanism in which its kinase activity cooperates with an adaptor function for EXOC to inhibit host cell actin dynamics.

IMPORTANCE
Human immunodeficiency virus type 1 (HIV-1) Nef contributes to AIDS pathogenesis, but the underlying molecular mechanisms remain incompletely understood. An important aspect of Nef function is to facilitate virus replication by disrupting T lymphocyte actin dynamics in response to stimulation via its association with the host cell kinase PAK2. We report here that the molecular surface of Nef for PAK2 association also mediates interaction of Nef with EXOC and establish that PAK2 provides an essential adaptor function for the subsequent formation of Nef-EXOC complexes. PAK2 and EXOC specifically cooperate in the inhibition of actin dynamics and proximal signaling induced by T cell receptor engagement by Nef. These results establish EXOC as a functionally relevant Nef interaction partner, emphasize the suitability of the PAK2 interaction surface for future therapeutic interference with Nef function, and show that such strategies need to target activity-independent PAK2 functions.
N
-Acetylglucosamine-Induced Cell Death in Candida albicans and Its Implications for Adaptive Mechanisms of Nutrient Sensing in Yeasts
Authors: Du Han , Guan Guobo , Li Xiaoling , Gulati Megha , Tao Li , Cao Chengjun , Johnson Alexander D. , Nobile Clarissa J. , Huang Guanghua
Published: September 2015

Single-celled organisms have different strategies to sense and utilize nutrients in their ever-changing environments. The opportunistic fungal pathogen
Candida albicans
is a common member of the human microbiota, especially that of the gastrointestinal (GI) tract. An important question concerns how
C. albicans
gained a competitive advantage over other microbes to become a successful commensal and opportunistic pathogen. Here, we report that
C. albicans
uses
N
-acetylglucosamine (GlcNAc), an abundant carbon source present in the GI tract, as a signal for nutrient availability. When placed in water,
C. albicans
cells normally enter the G
0
phase and remain viable for weeks. However, they quickly lose viability when cultured in water containing only GlcNAc. We term this phenomenon GlcNAc-induced cell death (GICD). GlcNAc triggers the upregulation of ribosomal biogenesis genes, alterations of mitochondrial metabolism, and the accumulation of reactive oxygen species (ROS), followed by rapid cell death via both apoptotic and necrotic mechanisms. Multiple pathways, including the conserved cyclic AMP (cAMP) signaling and GlcNAc catabolic pathways, are involved in GICD. GlcNAc acts as a signaling molecule to regulate multiple cellular programs in a coordinated manner and therefore maximizes the efficiency of nutrient use. This adaptive behavior allows
C. albicans
’ more efficient colonization of the gut.

IMPORTANCE
The ability to rapidly and appropriately respond to nutrients in the environment is crucial to free-living microorganisms. To maximize the use of available nutrients, microorganisms often use a limiting nutritional component as a signal to coordinate multiple biological processes. The human fungal pathogen
Candida albicans
uses
N
-acetylglucosamine (GlcNAc) as a signal for the availability of external nutrient resources. GlcNAc induces rapid cell death in
C. albicans
due to the constitutive activation of oxidative metabolism and accumulation of reactive oxygen species (ROS), and multiple pathways are involved in its regulation. This study sheds light on the mechanisms of niche specialization of pathogenic fungi and raises the possibility that this cell death pathway could be an unexplored therapeutic target.
Factors related to previous tuberculosis treatment of patients with multidrug-resistant tuberculosis in Bangladesh
Authors: Rifat Mahfuza , Hall John , Oldmeadow Christopher , Husain Ashaque , Hinderaker Sven Gudmund , Milton Abul Hasnat
Published: September 2015

Objective

Previous tuberculosis (TB) treatment status is an established risk factor for multidrug-resistant TB (MDR-TB). This study explores which factors related to previous TB treatment may lead to the development of multidrug resistant in Bangladesh.

Design

We previously conducted a large case–control study to identify risk factors for developing MDR-TB in Bangladesh. Patients who had a history of previous TB treatment, either MDR-TB or non-MDR-TB, were interviewed about their previous treatment episode. This study restricts analysis to the strata of patients who have been previously treated for TB. Information was collected through face-to-face interviews and record reviews. Unadjusted and multivariable logistic regression was used for data analysis.

Setting

Central-level, district-level and subdistrict-level hospitals in rural and urban Bangladesh.

Results

The strata of previously treated patients include a total of 293 patients (245 current MDR-TB; 48 non-MDR-TB). Overall, 54% of patients received previous TB treatment more than once, and all of these patients were multidrug resistant. Patients with MDR-TB were more likely to have experienced the following factors: incomplete treatment (OR 4.3; 95% CI 1.7 to 10.6), adverse reactions due to TB treatment (OR 8.2; 95% CI 3.2 to 20.7), hospitalisation for symptoms associated with TB (OR 16.9; CI 1.8 to 156.2), DOTS (directly observed treatment, short-course) centre as treatment unit (OR 6.4; CI 1.8 to 22.8), supervised treatment (OR 3.8; CI 1.6 to 9.5); time-to-treatment centre (OR 0.984; CI 0.974 to 0.993).

Conclusions

Incomplete treatment, hospitalisation for TB treatment and adverse reaction are the factors related to previous TB treatment of patients with MDR-TB. Although the presence of supervised treatment (DOT), less time-to-treatment centres and being treated in DOTS centres were relatively higher among the patients with MDR-TB compared with patients without MDR-TB, these findings include information of their most recent TB treatment episode only. Most (64.5%) of the patients with MDR-TB had received TB treatment more than once.
A retrospective pilot study of correlation of cerebral augmentation effects of external counterpulsation with functional outcome after acute ischaemic stroke
Authors: Xiong Li , Lin Wenhua , Han Jinghao , Chen Xiangyan , Leung Thomas Wai Hong , Soo Yannie Oi Yan , Wong Lawrence Ka Sing
Published: September 2015

Objective

External counterpulsation (ECP) is a non-invasive method used to augment cerebral blood flow of patients with ischaemic stroke via induced hypertension. We aimed to explore the correlation between the cerebral blood flow augmentation effects induced by ECP and clinical outcome after acute ischaemic stroke.

Methods

We retrospectively analysed our ECP registry of patients with ischaemic stroke who were enrolled within 7 days after stroke onset. Bilateral middle cerebral arteries of patients were monitored using transcranial Doppler (TCD). Flow velocity changes before, during and after ECP were, respectively, recorded for 3 min. The cerebral augmentation index (CAI) was the increase in percentage of the middle cerebral artery mean flow velocity during ECP compared with baseline. TCD data were analysed based on the side ipsilateral or contralateral to the infarct. The modified Rankin Scale (mRS) (good outcome: mRS 0∼2; poor outcome: mRS 3∼6) was evaluated 6 months after the index stroke.

Results

72 patients were included (mean age, 63.8±10.7 years; 87.5% males). At month 6 after stroke onset, univariate analysis showed that the National Institutes of Health Stroke Scale at recruitment was significantly higher and ECP therapy duration was longer in the poor outcome group, while the ipsilateral CAI was significantly lower in the good outcome group than that in the poor outcome group (3.71±4.94 vs 7.73±7.66, p=0.044). Multivariate logistic regression showed that ipsilateral CAI was independently correlated with an unfavourable functional outcome after adjusting for confounding factors.

Conclusions

The higher degree of cerebral blood flow velocity augmentation on the side ipsilateral to the infarct induced by ECP is independently correlated with an unfavourable functional outcome after acute ischaemic stroke.
Pre-trial inter-laboratory analytical validation of the FOCUS4 personalised therapy trial
Authors: Richman Susan D , Adams Richard , Quirke Phil , Butler Rachel , Hemmings Gemma , Chambers Phil , Roberts Helen , James Michelle D , Wozniak Sue , Bathia Riya , Pugh Cheryl , Maughan Timothy , Jasani Bharat
Published: September 2015

Introduction

Molecular characterisation of tumours is increasing personalisation of cancer therapy, tailored to an individual and their cancer. FOCUS4 is a molecularly stratified clinical trial for patients with advanced colorectal cancer. During an initial 16-week period of standard first-line chemotherapy, tumour tissue will undergo several molecular assays, with the results used for cohort allocation, then randomisation. Laboratories in Leeds and Cardiff will perform the molecular testing. The results of a rigorous pre-trial inter-laboratory analytical validation are presented and discussed.

Methods

Wales Cancer Bank supplied FFPE tumour blocks from 97 mCRC patients with consent for use in further research. Both laboratories processed each sample according to an agreed definitive FOCUS4 laboratory protocol, reporting results directly to the MRC Trial Management Group for independent cross-referencing.

Results

Pyrosequencing analysis of mutation status atKRAScodons12/13/61/146,NRAScodons12/13/61,BRAFcodon600 andPIK3CAcodons542/545/546/1047, generated highly concordant results. Two samples gave discrepant results; in one aPIK3CAmutation was detected only in Leeds, and in the other, aPIK3CAmutation was only detected in Cardiff. pTEN and mismatch repair (MMR) protein expression was assessed by immunohistochemistry (IHC) resulting in 6/97 discordant results for pTEN and 5/388 for MMR, resolved upon joint review. Tumour heterogeneity was likely responsible for pyrosequencing discrepancies. The presence of signet-ring cells, necrosis, mucin, edge-effects and over-counterstaining influenced IHC discrepancies.

Conclusions

Pre-trial assay analytical validation is essential to ensure appropriate selection of patients for targeted therapies. This is feasible for both mutation testing and immunohistochemical assays and must be built into the workup of such trials.

Trial registration number

ISRCTN90061564.