bluuebunny/crossref_metadata_2025 huggingface dataset:
Doubly Spliced RNA of Hepatitis B Virus Suppresses Viral Transcription via TATA-Binding Protein and Induces Stress Granule Assembly
Authors: Tsai Kuen-Nan , Chong Chin-Liew , Chou Yu-Chi , Huang Chien-Chiao , Wang Yi-Ling , Wang Shao-Win , Chen Mong-Liang , Chen Chun-Hong , Chang Chungming
Published: September 2015

The risk of liver cancer in patients infected with the hepatitis B virus (HBV) and their clinical response to interferon alpha therapy vary based on the HBV genotype. The mechanisms underlying these differences in HBV pathogenesis remain unclear. In HepG2 cells transfected with a mutant HBV
G2335A
expression plasmid that does not transcribe the 2.2-kb doubly spliced RNA (2.2DS-RNA) expressed by wild-type HBV genotype A, the level of HBV pregenomic RNA (pgRNA) was higher than that in cells transfected with an HBV genotype A expression plasmid. By using cotransfection with HBV genotype D and 2.2DS-RNA expression plasmids, we found that a reduction of pgRNA was observed in the cells even in the presence of small amounts of the 2.2DS-RNA plasmid. Moreover, ectopic expression of 2.2DS-RNA in the HBV-producing cell line 1.3ES2 reduced the expression of pgRNA. Further analysis showed that exogenously transcribed 2.2DS-RNA inhibited a reconstituted transcription
in vitro
. In Huh7 cells ectopically expressing 2.2DS-RNA, RNA immunoprecipitation revealed that 2.2DS-RNA interacted with the TATA-binding protein (TBP) and that nucleotides 432 to 832 of 2.2DS-RNA were required for efficient TBP binding. Immunofluorescence experiments showed that 2.2DS-RNA colocalized with cytoplasmic TBP and the stress granule components, G3BP and poly(A)-binding protein 1 (PABP1), in Huh7 cells. In conclusion, our study reveals that 2.2DS-RNA acts as a repressor of HBV transcription through an interaction with TBP that induces stress granule formation. The expression of 2.2DS-RNA may be one of the viral factors involved in viral replication, which may underlie differences in clinical outcomes of liver disease and responses to interferon alpha therapy between patients infected with different HBV genotypes.

IMPORTANCE
Patients infected with certain genotypes of HBV have a lower risk of hepatocellular carcinoma and exhibit a more favorable response to antiviral therapy than patients infected with other HBV genotypes. Using cultured human hepatoma cells as a model of HBV infection, we found that the expression of 2.2DS-RNA caused a decrease in HBV replication. In cultured cells, the ectopic expression of 2.2DS-RNA obviously reduced the intracellular levels of HBV mRNAs. Our analysis of the 2.2DS-RNA-mediated suppression of viral RNA expression showed that 2.2DS-RNA inhibited transcription via binding to the TATA-binding protein and stress granule proteins. Our findings suggest that the 2.2DS-RNA acts as a suppressive noncoding RNA that modulates HBV replication, which may in turn influence the development of chronic hepatitis B.
Signaling Lymphocytic Activation Molecule Family Receptor Homologs in New World Monkey Cytomegaloviruses
Authors: Pérez-Carmona Natàlia , Farré Domènec , Martínez-Vicente Pablo , Terhorst Cox , Engel Pablo , Angulo Ana
Published: September 2015

Throughout evolution, large DNA viruses have been usurping genes from their hosts to equip themselves with proteins that restrain host immune defenses. Signaling lymphocytic activation molecule (SLAM) family (SLAMF) receptors are involved in the regulation of both innate and adaptive immunity, which occurs upon engagement with their ligands via homotypic or heterotypic interactions. Here we report a total of seven SLAMF genes encoded by the genomes of two cytomegalovirus (CMV) species, squirrel monkey CMV (SMCMV) and owl monkey CMV (OMCMV), that infect New World monkeys. Our results indicate that host genes were captured by retrotranscription at different stages of the CMV-host coevolution. The most recent acquisition led to S1 in SMCMV. S1 is a SLAMF6 homolog with an amino acid sequence identity of 97% to SLAMF6 in its ligand-binding N-terminal Ig domain. We demonstrate that S1 is a cell surface glycoprotein capable of binding to host SLAMF6. Furthermore, the OMCMV genome encodes A33, an LY9 (SLAMF3) homolog, and A43, a CD48 (SLAMF2) homolog, two soluble glycoproteins which recognize their respective cellular counterreceptors and thus are likely to be viral SLAMF decoy receptors. In addition, distinct copies of further divergent CD48 homologs were found to be encoded by both CMV genomes. Remarkably, all these molecules display a number of unique features, including cytoplasmic tails lacking characteristic SLAMF signaling motifs. Taken together, our findings indicate a novel immune evasion mechanism in which incorporation of host SLAMF receptors that retain their ligand-binding properties enables viruses to interfere with SLAMF functions and to supply themselves with convenient structural molds for expanding their immunomodulatory repertoires.

IMPORTANCE
The way in which viruses shape their genomes under the continual selective pressure exerted by the host immune system is central for their survival. Here, we report that New World monkey cytomegaloviruses have broadly captured and duplicated immune cell receptors of the signaling lymphocyte activation molecule (SLAM) family during host-virus coevolution. Notably, we demonstrate that several of these viral SLAMs exhibit exceptional preservation of their N-terminal immunoglobulin domains, which results in maintenance of their ligand-binding capacities. At the same time, these molecules present distinctive structural properties which include soluble forms and the absence of typical SLAM signaling motifs in their cytoplasmic domains, likely reflecting the evolutionary adaptation undergone to efficiently interfere with host SLAM family activities. The observation that the genomes of other large DNA viruses might bear SLAM family homologs further underscores the importance of these molecules as a novel class of immune regulators and as convenient scaffolds for viral evolution.
Epstein-Barr Viruses (EBVs) Deficient in EBV-Encoded RNAs Have Higher Levels of Latent Membrane Protein 2 RNA Expression in Lymphoblastoid Cell Lines and Efficiently Establish Persistent Infections in Humanized Mice
Authors: Gregorovic Goran , Boulden Elizabeth A. , Bosshard Rachel , Elgueta Karstegl Claudio , Skalsky Rebecca , Cullen Bryan R. , Gujer Cornelia , Rämer Patrick , Münz Christian , Farrell Paul J.
Published: September 2015

Functions of Epstein-Barr virus (EBV)-encoded RNAs (EBERs) were tested in lymphoblastoid cell lines containing EBER mutants of EBV. Binding of EBER1 to ribosomal protein L22 (RPL22) was confirmed. Deletion of EBER1 or EBER2 correlated with increased levels of cytoplasmic EBV LMP2 RNA and with small effects on specific cellular microRNA (miRNA) levels, but protein levels of LMP1 and LMP2A were not affected. Wild-type EBV and EBER deletion EBV had approximately equal abilities to infect immunodeficient mice reconstituted with a human hematopoietic system.
Structural Basis of the Inhibition of STAT1 Activity by Sendai Virus C Protein
Authors: Oda Kosuke , Matoba Yasuyuki , Irie Takashi , Kawabata Ryoko , Fukushi Masaya , Sugiyama Masanori , Sakaguchi Takemasa
Published: September 2015

Sendai virus (SeV) C protein inhibits the signal transduction pathways of interferon alpha/beta (IFN-α/β) and IFN-γ by binding to the N-terminal domain of STAT1 (STAT1ND), thereby allowing SeV to escape from host innate immunity. Here we determined the crystal structure of STAT1ND associated with the C-terminal half of the C protein (Y3 [amino acids 99 to 204]) at a resolution of 2.0 Å. This showed that two molecules of Y3 symmetrically bind to each niche created between two molecules of the STAT1ND dimer. Molecular modeling suggested that an antiparallel form of the full-length STAT1 dimer can bind only one Y3 molecule and that a parallel form can bind two Y3 molecules. Affinity analysis demonstrated anticooperative binding of two Y3 molecules with the STAT1 dimer, which is consistent with the hypothetical model that the second Y3 molecule can only target the STAT1 dimer in a parallel form. STAT1 with excess amounts of Y3 was prone to inhibit the dephosphorylation at Tyr
701
by a phosphatase. In an electrophoretic mobility shift assay, tyrosine-phosphorylated STAT1 (pY-STAT1) with Y3 associated with the γ-activated sequence, probably as high-molecular-weight complexes (HMWCs), which may account for partial inhibition of a reporter assay from IFN-γ by Y3. Our study suggests that the full-length C protein interferes with the domain arrangement of the STAT1 dimer, leading to the accumulation of pY-STAT1 and the formation of HMWCs. In addition, we discuss the mechanism by which phosphorylation of STAT2 is inhibited in the presence of the C protein after stimulation by IFN-α/β.

IMPORTANCE
Sendai virus, a paramyxovirus that causes respiratory diseases in rodents, possesses the C protein, which inhibits the signal transduction pathways of interferon alpha/beta (IFN-α/β) and IFN-γ by binding to the transcription factor STAT1. In virus-infected cells, phosphorylation of STAT1 at the Tyr
701
residue is potently enhanced, although transcription by STAT1 is inert. Here, we determined the crystal structure of the N-terminal domain of STAT1 associated with the C-terminal half of the C protein. Molecular modeling and experiments suggested that the two C proteins bind to and stabilize the parallel form of the STAT1 dimer, which are likely to be phosphorylated at Tyr
701
, further inducing high-molecular-weight complex formation and inhibition of transcription by IFN-γ. We also discuss the possible mechanism of inhibition of the IFN-α/β pathways by the C protein. This is the first structural report of the C protein, suggesting a mechanism of evasion of the paramyxovirus from innate immunity.
The ns12.9 Accessory Protein of Human Coronavirus OC43 Is a Viroporin Involved in Virion Morphogenesis and Pathogenesis
Authors: Zhang Ronghua , Wang Kai , Ping Xianqiang , Yu Wenjing , Qian Zhikang , Xiong Sidong , Sun Bing
Published: September 2015

An accessory gene between the S and E gene loci is contained in all coronaviruses (CoVs), and its function has been studied in some coronaviruses. This gene locus in human coronavirus OC43 (HCoV-OC43) encodes the ns12.9 accessory protein; however, its function during viral infection remains unknown. Here, we engineered a recombinant mutant virus lacking the ns12.9 protein (HCoV-OC43-Δns12.9) to characterize the contributions of ns12.9 in HCoV-OC43 replication. The ns12.9 accessory protein is a transmembrane protein and forms ion channels in bothXenopusoocytes and yeast through homo-oligomerization, suggesting that ns12.9 is a newly recognized viroporin. HCoV-OC43-Δns12.9 presented at least 10-fold reduction of viral titerin vitroandin vivo. Intriguingly, exogenous ns12.9 and heterologous viroporins with ion channel activity could compensate for the production of HCoV-OC43-Δns12.9, indicating that the ion channel activity of ns12.9 plays a significant role in the production of infectious virions. Systematic dissection of single-cycle replication revealed that ns12.9 protein had no measurable effect on virus entry, subgenomic mRNA (sgmRNA) synthesis, and protein expression. Further characterization revealed that HCoV-OC43-Δns12.9 was less efficient in virion morphogenesis than recombinant wild-type virus (HCoV-OC43-WT). Moreover, reduced viral replication, inflammatory response, and virulence in HCoV-OC43-Δns12.9-infected mice were observed compared to the levels for HCoV-OC43-WT-infected mice. Taken together, our results demonstrated that the ns12.9 accessory protein functions as a viroporin and is involved in virion morphogenesis and the pathogenesis of HCoV-OC43 infection.

IMPORTANCEHCoV-OC43 was isolated in the 1960s and is a major agent of the common cold. The functions of HCoV-OC43 structural proteins have been well studied, but few studies have focused on its accessory proteins. In the present study, we demonstrated that the ns12.9 protein is a newly recognized viroporin, and the ns12.9 gene knockout virus (HCoV-OC43-Δns12.9) presents a growth defectin vitroandin vivo. We identified the important functions of the ns12.9 viroporin in virion morphogenesis during HCoV-OC43 infection. Furthermore, mice infected with HCoV-OC43-Δns12.9 exhibited reduced inflammation and virulence accompanied by a lower titer in the brain than that of wild-type-infected mice, suggesting the ns12.9 viroporin influences virus pathogenesis. Therefore, our findings revealed that the ns12.9 viroporin facilitates virion morphogenesis to enhance viral production, and these results provided a deeper understanding of HCoV-OC43 pathogenesis.
STAT3 Regulates Lytic Activation of Kaposi's Sarcoma-Associated Herpesvirus
Authors: King Christine A. , Li Xiaofan , Barbachano-Guerrero Arturo , Bhaduri-McIntosh Sumita
Published: September 2015

Lytic activation of Kaposi's sarcoma-associated herpesvirus (KSHV) from latency is a critical contributor to pathogenesis and progression of KSHV-mediated disease. Development of targeted treatment strategies and improvement of lytic-phase-directed oncolytic therapies, therefore, hinge on gaining a better understanding of latency-to-lytic-phase transition. A key observation in that regard, also common to other herpesviruses, is the partial permissiveness of latently infected cells to lytic-cycle-inducing agents. Here, we address the molecular basis of why only some KSHV-infected cells respond to lytic stimuli. Since cellular signal transducer and activator of transcription 3 (STAT3) is constitutively active in KSHV-associated cancers, KSHV activates STAT3, and STAT3 has been found to regulate lytic activation of Epstein-Barr virus (EBV)-infected cells, we asked if STAT3 contributes similarly to the life cycle of KSHV. We found that high levels of STAT3 correlate with the refractory state at the single-cell level under conditions of both spontaneous and induced lytic activation; importantly, STAT3 also regulates lytic susceptibility. Further, knockdown of STAT3 suppresses the cellular transcriptional corepressor Krüppel-associated box domain-associated protein 1 (KAP1; also known as TRIM28), and suppression of KAP1 activates lytic genes, including the viral lytic switch RTA, thereby linking STAT3 via KAP1 to regulation of the balance between lytic and latent cells. These findings, taken together with those from EBV-infected and, more recently, herpes simplex virus 1 (HSV-1)-infected cells, cement the contribution of host STAT3 to persistence of herpesviruses and simultaneously reveal an important lead to devise strategies to improve lytic-phase-directed therapies for herpesviruses.

IMPORTANCELytic activation of the cancer-causing Kaposi's sarcoma-associated herpesvirus (KSHV) is vital to its life cycle and causation of disease. Like other herpesviruses, however, a substantial fraction of latently infected cells are resistant to lytic-phase-inducing stimuli. Investigating the molecular basis for this refractory state is essential for understanding how the virus persists and how it causes disease and to guide efforts to improve treatment of KSHV-mediated diseases. We found that, like two other herpesviruses, EBV and HSV-1, KSHV exploits the cellular transcription factor STAT3 to regulate the susceptibility of latently infected cells to lytic triggers. These findings highlight a common STAT3-centered strategy used by herpesviruses to maintain persistence in their hosts while also revealing a key molecule to pursue while devising methods to improve herpesvirus lytic-phase-directed therapies.
Downregulation of Toll-Like Receptor 9 Expression by Beta Human Papillomavirus 38 and Implications for Cell Cycle Control
Authors: Pacini Laura , Savini Claudia , Ghittoni Raffaella , Saidj Djamel , Lamartine Jerome , Hasan Uzma A. , Accardi Rosita , Tommasino Massimo
Published: September 2015

Innate immunity is the first line of host defense against infections. Many oncogenic viruses can deregulate several immune-related pathways to guarantee the persistence of the infection. Here, we show that the cutaneous human papillomavirus 38 (HPV38) E6 and E7 oncoproteins suppress the expression of the double-stranded DNA sensor Toll-like receptor 9 (TLR9) in human foreskin keratinocytes (HFK), a key mediator of the antiviral innate immune host response. In particular, HPV38 E7 induces TLR9 mRNA downregulation by promoting accumulation of ΔNp73α, an antagonist of p53 and p73. Inhibition of ΔNp73α expression by antisense oligonucleotide in HPV38 E6/E7 HFK strongly rescues mRNA levels of TLR9, highlighting a key role of ΔNp73α in this event. Chromatin immunoprecipitation experiments showed that ΔNp73α is part of a negative transcriptional regulatory complex with IκB kinase beta (IKKβ) that binds to a NF-κB responsive element within the TLR9 promoter. In addition, the Polycomb protein enhancer of zeste homolog 2 (EZH2), responsible for gene expression silencing, is also recruited into the complex, leading to histone 3 trimethylation at lysine 27 (H3K27me3) in the same region of the TLR9 promoter. Ectopic expression of TLR9 in HPV38 E6/E7 cells resulted in an accumulation of the cell cycle inhibitors p21
WAF1
and p27
Kip1
, decreased CDK2-associated kinase activity, and inhibition of cellular proliferation. In summary, our data show that HPV38, similarly to other viruses with well-known oncogenic activity, can downregulate TLR9 expression. In addition, they highlight a new role for TLR9 in cell cycle regulation.

IMPORTANCE
The mucosal high-risk HPV types have been clearly associated with human carcinogenesis. Emerging lines of evidence suggest the involvement of certain cutaneous HPV types in development of skin squamous cell carcinoma, although this association is still under debate. Oncogenic viruses have evolved different strategies to hijack the host immune system in order to guarantee the persistence of the infection. Their capability to evade the immune system is as important as their ability to promote cellular transformation. Therefore, understanding the viral mechanisms involved in viral persistence is a valid tool to evaluate their potential role in human carcinogenesis. Here, we show that E6 and E7 oncoproteins from the cutaneous HPV38 downregulate the expression of the double-stranded DNA sensor TLR9 of innate immunity. We also present evidence that the HPV38-mediated downregulation of TLR9 expression, in addition to its potential impact on the innate immune response, is linked to cell cycle deregulation.
Neuromyelitis optica and myasthenia gravis in a young Nigerian girl
Authors: Balarabe Salisu Abdullahi , Adamu Mohammad Dantani , Watila Musa Mamman , Jiya Nma
Published: September 2015

Neuromyelitis optica (NMO) and myasthenia gravis (MG) are rare autoimmune disorders. The coexistence of the two disorders, although rare, has been documented. This is a case report of a 16-year-old student who presented with recurrent episodes of transverse myelitis and optic neuritis, 8 years after diagnosis of MG. She presented with visual impairment, relapsing and remitting weakness, numbness and paraesthesia of her lower limbs, with bladder and bowel incontinence. Her examination revealed bilateral optic atrophy, spastic paraparesis of the lower limbs and patchy sensory loss up to thoracic level (T4-5). She had a positive acetylcholine receptor antibody, a positive aquaporin-4 antibody and chest CT finding of thymic enlargement. We therefore confirmed the previous diagnosis of MG and performed a recent diagnosis of background NMO. A high index of suspicion is needed to make a diagnosis of this rare coexistence of NMO and MG in resource-limited settings such as Nigeria.
Primary bone metastasis as first manifestation of an unknown primary tumour
Authors: García Carretero Rafael , Romero Brugera Marta , Rebollo-Aparicio Noelia , El Bouayadi Mohamed Liliam
Published: September 2015

Unknown primary tumour refers to a group of cancers for which the anatomical site of origin remains occult after detailed investigations. Thanks to sophisticated imaging, immunohistochemical testing and molecular-profiling tools, there is a more accurate approach to unknown primary cancer. Metastasis to bone is not a rare phenomenon, because any tumour can metastasise to bone, so it is a common clinical scenario. The role of clinical history, physical examination, laboratory tests, radiographic studies and immunohistochemistry is critical for a successful diagnostic strategy. Subsets of unknown primary cancers can be identified primarily on the basis of histopathological findings, the pattern of spread and serum markers. New immunohistochemical markers and tissue-of-origin profiles may establish presumable primary sites to unknown primary cancer on the basis of immunohistochemical and molecular patterns. We present a case of a 57-year-old woman without a cancer history, who had primary bone metastasis as the first manifestation of an occult primary tumour.
Variation in hospital rates of induction of labour: a population-based record linkage study
Authors: Nippita Tanya A , Trevena Judy A , Patterson Jillian A , Ford Jane B , Morris Jonathan M , Roberts Christine L
Published: September 2015

Objectives

To examine interhospital variation in rates of induction of labour (IOL) to identify potential targets to reduce high rates of practice variation.

Design

Population-based record linkage cohort study.

Setting

New South Wales, Australia, 2010–2011.

Participants

All women with live births of ≥24 weeks gestation in 72 hospitals.

Primary outcome measure

Variation in hospital IOL rates adjusted for differences in case-mix, according to 10 mutually exclusive groups derived from the Robson caesarean section classification; groups were categorised by parity, plurality, fetal presentation, prior caesarean section and gestational age.

Results

The overall IOL rate was 26.7% (46 922 of 175 444 maternities were induced), ranging from 9.7% to 41.2% (IQR 21.8–29.8%) between hospitals. Nulliparous and multiparous women at 39–40 weeks gestation with a singleton cephalic birth were the greatest contributors to the overall IOL rate (23.5% and 20.2% of all IOL respectively), and had persisting high unexplained variation after adjustment for case-mix (adjusted hospital IOL rates ranging from 11.8% to 44.9% and 7.1% to 40.5%, respectively). In contrast, there was little variation in interhospital IOL rates among multiparous women with a singleton cephalic birth at ≥41 weeks gestation, women with singleton non-cephalic pregnancies and women with multifetal pregnancies.

Conclusions

7 of the 10 groups showed high or moderate unexplained variation in interhospital IOL rates, most pronounced for women at 39–40 weeks gestation with a singleton cephalic birth. Outcomes associated with divergent practice require determination, which may guide strategies to reduce practice variation.
Atrial fibrillation care improvement collaborative
Authors: Robelia Paul , Kopecky Stephen , Thacher Tom
Published: September 2015

Atrial fibrillation (AF) is an increasingly common cardiac arrhythmia. Many patients with new onset or recurrent AF present to the emergency department and are subsequently admitted to the hospital and seen by cardiology specialists for follow up. In an attempt to address this high utilization of acute health care resources, reduce costs, and improve patient care, our institution instituted a collaborative project between the departments of emergency medicine, cardiology, family medicine, and primary care internal medicine.

The project team oversaw development of a new emergency department AF order set, encouraged utilization of a new oral anticoagulant (dabigatran), improved the primary care follow up connection, and deployed a multimodal education plan for primary care providers. Between 2012 and 2014, these interventions resulted in a 17% reduction in total AF per member per month (PMPM) cost, a 28% reduction in AF PMPM inpatient cost, and a 24% reduction in inpatient admissions for AF.
Compliance to two city convenience store ordinance requirements
Authors: Chaumont Menéndez Cammie K , Amandus Harlan E , Wu Nan , Hendricks Scott A
Published: September 2015

Background

Robbery-related homicides and assaults are the leading cause of death in retail businesses. Robbery reduction approaches focus on compliance to Crime Prevention Through Environmental Design (CPTED) guidelines.

Purpose

We evaluated the level of compliance to CPTED guidelines specified by convenience store safety ordinances effective in 2010 in Dallas and Houston, Texas, USA.

Methods

Convenience stores were defined as businesses less than 10 000 square feet that sell grocery items. Store managers were interviewed for store ordinance requirements from August to November 2011, in a random sample of 594 (289 in Dallas, 305 in Houston) convenience stores that were open before and after the effective dates of their city's ordinance. Data were collected in 2011 and analysed in 2012–2014.

Results

Overall, 9% of stores were in full compliance, although 79% reported being registered with the police departments as compliant. Compliance was consistently significantly higher in Dallas than in Houston for many requirements and by store type. Compliance was lower among single owner-operator stores compared with corporate/franchise stores. Compliance to individual requirements was lowest for signage and visibility.

Conclusions

Full compliance to the required safety measures is consistent with industry ‘best practices’ and evidence-based workplace violence prevention research findings. In Houston and Dallas compliance was higher for some CPTED requirements but not the less costly approaches that are also the more straightforward to adopt.
‘Off colour’ alpaca
Authors: Turner Andrea
Published: September 2015

THIS series gives readers the opportunity to consider and contribute to discussion of some of the ethical dilemmas that can arise in veterinary practice. Each month, a case scenario is presented, followed by discussion of some of the issues involved.

In addition, a possible way forward is suggested; however, there is rarely a cut‐and‐dried answer in such cases, and readers may wish to suggest an alternative approach. This month's dilemma, ‘“Off colour” alpaca?’, was submitted and is discussed by Andrea Turner. Readers with comments to contribute are invited to send them as soon as possible, so that they can be considered for publication in the next issue. Discussion of the dilemma ‘Billy the kid’, which was published in the July/August issue of In Practice, appears on page 431.

The series is being coordinated by Siobhan Mullan, of the University of Bristol. It is hoped it will provide a framework that will help practices find solutions when facing similar dilemmas.
Management of gundog injuries
Authors: Houlton John
Published: September 2015

Gundogs suffer from numerous injuries, many of which are minor and treated by their owners or handlers. These dogs undertake a diversity of work rather than having a single role, so when veterinary treatment is sought advice should be tailored accordingly; owners will also be anxious to minimise the time off work during the UK gamebird shooting season. This article describes the types of injuries sustained by gundogs and how they can be managed, taking into account the role of the dog.
Republished: Treatment of post-traumatic carotid–cavernous fistulas using pipeline embolization device assistance
Authors: Pradeep Nisha , Nottingham Robert , Kam Anthony , Gandhi Dheeraj , Razack Nasser
Published: September 2015

This report describes two cases of post-traumatic, high flow carotid–cavernous fistulas that demonstrated residual shunting after initial embolization with coils and Onyx, and that were successfully closed with pipeline embolization devices. Following their combined endovascular treatments, the patients experienced clinical improvement of symptoms with durable obliteration of the fistulous communications.
Specificity of the comparative skin test for bovine tuberculosis in Great Britain
Authors: Goodchild A. V. , Downs S. H. , Upton P. , Wood J. L. N. , de la Rua‐Domenech R.
Published: September 2015

A novel method for estimating specificity of the Single Intradermal Comparative Cervical Tuberculin (SICCT) test for bovine tuberculosis (bTB) using surveillance tests results is reported. The specificity of the SICCT test at three cut‐offs was estimated from the dates, locations and skinfold measurements of all routine tests carried out in Officially TB Free (OTF) cattle herds in Great Britain (GB) between 2002 and 2008, according to their separation (by distance and time) from known infected (OTF‐withdrawn) herds. The proportion of animals that tested positive was constant (P>0.20) when the distance between tested herds and nearest infected herd exceeded 8 km. For standard cut‐off, calculated specificity was 99.98 per cent (95 per cent confidence interval ±0.004 per cent), equating to one false positive result per 5000 uninfected animals tested. For severe cut‐off it was 99.91 per cent (±0.013 per cent) and for ultrasevere cut‐off (selecting all reactors and inconclusive reactors) it was 99.87 per cent (±0.017 per cent). The estimated positive predictive value of the test averaged 91 per cent and varied by regional prevalence. This study provides further evidence of the high specificity of the SICCT test under GB conditions, suggests that over 90 per cent of cattle currently culled using this test in GB were infected, and endorses slaughter of at least these cattle for bTB control.
Estimating walleye (Sander vitreus) movement and fishing mortality using state-space models: implications for management of spatially structured populations
Authors: Herbst Seth J. , Stevens Bryan S. , Hayes Daniel B. , Hanchin Patrick A.
Published: September 2015

Fish often exhibit complex movement patterns, and quantification of these patterns is critical for understanding many facets of fisheries ecology and management. In this study, we estimated movement and fishing mortality rates for exploited walleye (Sander vitreus) populations in a lake-chain system in northern Michigan. We developed a state-space model to estimate lake-specific movement and fishery parameters and fit models to observed angler tag return data using Bayesian estimation and inference procedures. Informative prior distributions for lake-specific spawning-site fidelity, fishing mortality, and system-wide tag reporting rates were developed using auxiliary data to aid model-fitting. Our results indicated that postspawn movement among lakes was asymmetrical and ranged from approximately 1% to 42% per year, with the largest outmigration occurring from the Black River, which was primarily used by adult fish during the spawning season. Instantaneous fishing mortality rates differed among lakes and ranged from 0.16 to 0.27, with the highest rate coming from one of the smaller and uppermost lakes in the system. The approach developed provides a flexible framework that incorporates seasonal behavioral ecology (i.e., spawning-site fidelity) in estimation of movement for a mobile fish species that will ultimately provide information to aid research and management for spatially structured fish populations.
Recent advances in statistical methodology applied to the Hjort liver index time series (1859–2012) and associated influential factors
Authors: Hermansen Gudmund H. , Hjort Nils Lid , Kjesbu Olav S.
Published: September 2015

Certain recent advances in statistical methodology have promising potential for fruitful use in general biology and the fisheries sciences. This paper reviews and discusses some of the relevant themes, including accurate modelling via focused model selection techniques, dynamic goodness-of-fit testing of processes evolving over time, finding break points for phenomena experiencing regime shifts, prediction uncertainty, and optimal combination of information across diverse sources via confidence distributions. The methods are illustrated for the Hjort liver quality index time series. Its roots lie in the classic study by Hjort in 1914, where liver quality of the Northeast Arctic cod (Gadus morhua) for 1880–1912 was reported on and studied, along with related factors, making it one of the first teleost time series ever published. The series has been extended both backwards and forwards in time, to 1859–2012, due to comprehensive archival and calibration efforts of Kjesbu et al. in 2014, yielding one of the longest time series of marine science. Our study offers a detailed examination of this series and how it relates to and interacts with associated factors, including winter temperatures, length distribution parameters, cod mortality, and a certain index related to availability of food. We identify certain mild nonstationary aspects of the time series, show that there is a regime shift around 1990 in the ways the liver series interacts with winter temperatures, and demonstrate that mortality and food availability play important roles.
Factors controlling the abundance of rainbow trout in the Colorado River in Grand Canyon in a reach utilized by endangered humpback chub
Authors: Korman Josh , Yard Michael D. , Yackulic Charles B.
Published: September 2015

We estimated the abundance, survival, movement, and recruitment of non-native rainbow trout (Oncorhynchus mykiss) in the Colorado River in Grand Canyon to determine what controls their abundance near the Little Colorado River (LCR) confluence where endangered humpback chub (Gila cypha) rear. Over a 3-year period, we tagged more than 70 000 trout and recovered over 8200 tagged fish. Trout density was highest (10 000–25 000 fish·km–1) in the reach closest to Glen Canyon Dam, where the majority of trout recruitment occurs, and was 30- to 50-fold lower (200–800 fish·km–1) in reaches near the LCR confluence ∼100 km downstream. The extent of rainbow trout movement was limited with less than 1% of recaptures making movements greater than 20 km. However, because of high trout densities in upstream source areas, this small dispersal rate was sufficient to explain the threefold increase in the relatively small population near the LCR. Reducing dispersal rates of trout from upstream sources is the most feasible solution to maintain low densities near the LCR to minimize negative effects of competition and predation on humpback chub.
Stochastic Perturbations and Invariant Measures of Position Dependent Random Maps via Fourier Approximations
Authors: Islam Md Shafiqul
Published: September 2015

Let T = {τ1(x), τ2(x),…, τK(x); p1(x), p2(x),…, pK(x)} be a position dependent random map which possesses a unique absolutely continuous invariant measure [Formula: see text] with probability density function [Formula: see text]. We consider a family {TN}N≥1 of stochastic perturbations TN of the random map T. Each TN is a Markov process with the transition density [Formula: see text], where qN(x, ⋅) is a doubly stochastic periodic and separable kernel. Using Fourier approximation, we construct a finite dimensional approximation PN to a perturbed Perron–Frobenius operator. Let [Formula: see text] be a fixed point of PN. We show that [Formula: see text] converges in L1 to [Formula: see text].
Delay-Induced Bogdanov–Takens Bifurcation and Dynamical Classifications in a Slow-Fast Flexible Joint System
Authors: Xu Jian , Jiang Shanying
Published: September 2015

A slow-fast delay-coupled flexible joint system is investigated in this paper. To understand the effects of time delay on the stability and oscillation of the manipulator, the geometric singular perturbation method is extended in dealing with delay differential equations. Bogdanov–Takens (BT) bifurcation of the fast subsystem is obtained, which leads to the existence of homoclinic orbits and is proved to be related to the formation of spiking. After the break of homoclinic orbits, Melnikov theory is introduced to predict the threshold curve indicating the occurrence of chaos. Numerical results show that with the increase of time delay, the stability of the system gets worse, and complicated oscillations including bursting, chaotic-bursting and complete chaos turn up. Besides, it is briefly summarized that the effect of the small parameter in the slow-fast system is to influence the convergence rate of solution trajectories, which is widely neglected in previous works.
Activation of protein kinase Cα increases phosphorylation of the UT-A1 urea transporter at serine 494 in the inner medullary collecting duct
Authors: Blount Mitsi A. , Cipriani Penelope , Redd Sara K. , Ordas Ronald J. , Black Lauren N. , Gumina Diane L. , Hoban Carol A. , Klein Janet D. , Sands Jeff M.
Published: September 2015

Hypertonicity increases urea transport, as well as the phosphorylation and membrane accumulation of UT-A1, the transporter responsible for urea permeability in the inner medullary collect duct (IMCD). Hypertonicity stimulates urea transport through PKC-mediated phosphorylation. To determine whether PKC phosphorylates UT-A1, eight potential PKC phosphorylation sites were individually replaced with alanine and subsequently transfected into LLC-PK1 cells. Of the single mutants, only ablation of the S494 site dampened induction of total UT-A1 phosphorylation by the PKC activator phorbol dibutyrate (PDBu). This result was confirmed using a newly generated antibody that specifically detected phosphorylation of UT-A1 at S494. Hypertonicity increased UT-A1 phosphorylation at S494. In contrast, activators of cAMP pathways (PKA and Epac) did not increase UT-A1 phosphorylation at S494. Activation of both PKC and PKA pathways increased plasma membrane accumulation of UT-A1, although activation of PKC alone did not do so. However, ablating the PKC site S494 decreased UT-A1 abundance in the plasma membrane. This suggests that the cAMP pathway promotes UT-A1 trafficking to the apical membrane where the PKC pathway can phosphorylate the transporter, resulting in increased UT-A1 retention at the apical membrane. In summary, activation of PKC increases the phosphorylation of UT-A1 at a specific residue, S494. Although there is no cross talk with the cAMP-signaling pathway, phosphorylation of S494 through PKC may enhance vasopressin-stimulated urea permeability by retaining UT-A1 in the plasma membrane.
A practical method for monitoring FRET-based biosensors in living animals using two-photon microscopy
Authors: Tao Wen , Rubart Michael , Ryan Jennifer , Xiao Xiao , Qiao Chunping , Hato Takashi , Davidson Michael W. , Dunn Kenneth W. , Day Richard N.
Published: September 2015

The commercial availability of multiphoton microscope systems has nurtured the growth of intravital microscopy as a powerful technique for evaluating cell biology in the relevant context of living animals. In parallel, new fluorescent protein (FP) biosensors have become available that enable studies of the function of a wide range of proteins in living cells. Biosensor probes that exploit Förster resonance energy transfer (FRET) are among the most sensitive indicators of an array of cellular processes. However, differences between one-photon and two-photon excitation (2PE) microscopy are such that measuring FRET by 2PE in the intravital setting remains challenging. Here, we describe an approach that simplifies the use of FRET-based biosensors in intravital 2PE microscopy. Based on a systematic comparison of many different FPs, we identified the monomeric (m) FPs mTurquoise and mVenus as particularly well suited for intravital 2PE FRET studies, enabling the ratiometric measurements from linked FRET probes using a pair of experimental images collected simultaneously. The behavior of the FPs is validated by fluorescence lifetime and sensitized emission measurements of a set of FRET standards. The approach is demonstrated using a modified version of the AKAR protein kinase A biosensor, first in cells in culture, and then in hepatocytes in the liver of living mice. The approach is compatible with the most common 2PE microscope configurations and should be applicable to a variety of different FRET probes.
Rab-GAP TBC1D4 (AS160) is dispensable for the renal control of sodium and water homeostasis but regulates GLUT4 in mouse kidney
Authors: Di Chiara Marianna , Glaudemans Bob , Loffing-Cueni Dominique , Odermatt Alex , Al-Hasani Hadi , Devuyst Olivier , Faresse Nourdine , Loffing Johannes
Published: September 2015

The Rab GTPase-activating protein TBC1D4 (AS160) controls trafficking of the glucose transporter GLUT4 in adipocytes and skeletal muscle cells. TBC1D4 is also highly abundant in the renal distal tubule, although its role in this tubule is so far unknown. In vitro studies suggest that it is involved in the regulation of renal transporters and channels such as the epithelial sodium channel (ENaC), aquaporin-2 (AQP2), and the Na+-K+-ATPase. To assess the physiological role of TBC1D4 in the kidney, wild-type (TBC1D4+/+) and TBC1D4-deficient (TBC1D4−/−) mice were studied. Unexpectedly, neither under standard nor under challenging conditions (low Na+/high K+, water restriction) did TBC1D4−/−mice show any difference in urinary Na+and K+excretion, urine osmolarity, plasma ion and aldosterone levels, and blood pressure compared with TBC1D4+/+mice. Also, immunoblotting did not reveal any change in the abundance of major renal sodium- and water-transporting proteins [Na-K-2Cl cotransporter (NKCC2) NKCC2, NaCl cotransporter (NCC), ENaC, AQP2, and the Na+-K+-ATPase]. However, the abundance of GLUT4, which colocalizes with TBC1D4 along the distal nephron of TBC1D4+/+mice, was lower in whole kidney lysates of TBC1D4−/−mice than in TBC1D4+/+mice. Likewise, primary thick ascending limb (TAL) cells isolated from TBC1D4−/−mice showed an increased basal glucose uptake and an abrogated insulin response compared with TAL cells from TBC1D4+/+mice. Thus, TBC1D4 is dispensable for the regulation of renal Na+and water transport, but may play a role for GLUT4-mediated basolateral glucose uptake in distal tubules. The latter may contribute to the known anaerobic glycolytic capacity of distal tubules during renal ischemia.
Polarized ATP distribution in urothelial mucosal and serosal space is differentially regulated by stretch and ectonucleotidases
Authors: Yu Weiqun
Published: September 2015

Purinergic signaling is a major pathway in regulating bladder function, and mechanical force stimulates urothelial ATP release, which plays an important role in bladder mechanotransduction. Although urothelial ATP release was first reported almost 20 years ago, the way in which release is regulated by mechanical force, and the presence of ATP-converting enzymes in regulating the availability of released ATP is still not well understood. Using a set of custom-designed Ussing chambers with the ability to manipulate mechanical forces applied on the urothelial tissue, we have demonstrated that it is stretch and not hydrostatic pressure that induces urothelial ATP release. The experiments reveal that urothelial ATP release is tightly controlled by stretch speed, magnitude, and direction. We have further shown that stretch-induced urothelial ATP release is insensitive to temperature (4°C). Interestingly, stretch-induced ATP release shows polarized distribution, with the ATP concentration in mucosal chamber (nanomolar level) about 10 times higher than the ATP concentration in serosal chamber (subnanomolar level). Furthermore, we have consistently observed differential ATP lifetime kinetics in the mucosal and serosal chambers, which is consistent with our immunofluorescent localization data, showing that ATP-converting enzymes ENTPD3 and alkaline phosphatase are expressed on urothelial basal surface, but not on the apical membrane. In summary, our data indicate that urothelial ATP release is finely regulated by stretch speed, magnitude, and direction, and extracellular ATP signaling is likely to be differentially regulated by ectonucleotidase, which results in temporally and spatially distinct ATP kinetics in response to mechanical stretch.
Less pronounced response to exercise in healthy relatives to type 2 diabetic subjects compared with controls
Authors: Ekman C. , Elgzyri T. , Ström K. , Almgren P. , Parikh H. , Dekker Nitert Marloes , Rönn T. , Manderson Koivula Fiona , Ling C. , Tornberg Å. B. , Wollmer P. , Eriksson K. F. , Groop L. , Hansson O.
Published: September 2015

Healthy first-degree relatives with heredity of type 2 diabetes (FH+) are known to have metabolic inflexibility compared with subjects without heredity for diabetes (FH−). In this study, we aimed to test the hypothesis that FH+ individuals have an impaired response to exercise compared with FH−. Sixteen FH+ and 19 FH− insulin-sensitive men similar in age, peak oxygen consumption (V̇o2 peak), and body mass index completed an exercise intervention with heart rate monitored during exercise for 7 mo. Before and after the exercise intervention, the participants underwent a physical examination and tests for glucose tolerance and exercise capacity, and muscle biopsies were taken for expression analysis. The participants attended, on average, 39 training sessions during the intervention and spent 18.8 MJ on exercise. V̇o2 peak/kg increased by 14%, and the participants lost 1.2 kg of weight and 3 cm waist circumference. Given that the FH+ group expended 61% more energy during the intervention, we used regression analysis to analyze the response in the FH+ and FH− groups separately. Exercise volume had a significant effect on V̇o2 peak, weight, and waist circumference in the FH− group, but not in the FH+ group. After exercise, expression of genes involved in metabolism, oxidative phosphorylation, and cellular respiration increased more in the FH− compared with the FH+ group. This suggests that healthy, insulin-sensitive FH+ and FH− participants with similar age, V̇o2 peak, and body mass index may respond differently to an exercise intervention. The FH+ background might limit muscle adaptation to exercise, which may contribute to the increased susceptibility to type 2 diabetes in FH+ individuals.
Functional segregation of retinal ganglion cell projections to the optic tectum of rainbow trout
Authors: Novales Flamarique Iñigo , Wachowiak Matt
Published: September 2015

The interpretation of visual information relies on precise maps of retinal representation in the brain coupled with local circuitry that encodes specific features of the visual scenery. In nonmammalian vertebrates, the main target of ganglion cell projections is the optic tectum. Although the topography of retinotectal projections has been documented for several species, the spatiotemporal patterns of activity and how these depend on background adaptation have not been explored. In this study, we used a combination of electrical and optical recordings to reveal a retinotectal map of ganglion cell projections to the optic tectum of rainbow trout and characterized the spatial and chromatic distribution of ganglion cell fibers coding for increments (ON) and decrements (OFF) of light. Recordings of optic nerve activity under various adapting light backgrounds, which isolated the input of different cone mechanisms, yielded dynamic patterns of ON and OFF input characterized by segregation of these two fiber types. Chromatic adaptation decreased the sensitivity and response latency of affected cone mechanisms, revealing their variable contributions to the ON and OFF responses. Our experiments further demonstrated restricted input from a UV cone mechanism to the anterolateral optic tectum, in accordance with the limited presence of UV cones in the dorsotemporal retina of juvenile rainbow trout. Together, our findings show that retinal inputs to the optic tectum of this species are not homogeneous, exhibit highly dynamic activity patterns, and are likely determined by a combination of biased projections and specific retinal cell distributions and their activity states.