bluuebunny/crossref_metadata_2025 huggingface dataset:
Application of Schwarz–Christoffel mapping to the analysis of conduction through a slot
Authors: Yariv Ehud , Sherwood John D.
Published: August 2015

We consider the generic problem of steady conduction through a slot traversing a non-conducting plate that separates two semi-infinite conducting regions. The current-density field is conservative; the dimensionless problem governing its potential depends upon a single geometric parameter,
h
, the ratio of the slot length (i.e. the plate thickness) to its width. We construct a Schwarz–Christoffel transformation to handle this two-dimensional transport problem. The transformation is expressed in terms of two parameters which are related to
h
through two implicit equations; in the limit
h
→0, it becomes explicit. Because of the slow decay of the current density at large distances from the slot, the integral representing the slot resistance diverges. The excess resistance of a finite-length slot relative to that of a zero-length slot is, however, finite. This excess resistance depends only upon the asymptotic behaviour of the potential far from the slot; it may therefore be directly obtained as a function of the two transformation parameters. Asymptotic approximations are found for the excess resistance at small and large
h
, respectively, scaling as

h
ln
⁡
h

and
h
. The single-slot solution is used to analyse conduction through a periodic array of widely spaced slots.
The evolutionary puzzle of egg size, oxygenation and parental care in aquatic environments
Authors: Braga Goncalves Ines , Ahnesjö Ingrid , Kvarnemo Charlotta
Published: August 2015

Offspring fitness generally improves with increasing egg size. Yet, eggs of most aquatic organisms are small. A common but largely untested assumption is that larger embryos require more oxygen than they can acquire through diffusion via the egg surface, constraining egg size evolution. However, we found no detrimental effects of large egg size on embryo growth and survival under hypoxic conditions. We tested this in the broad-nosed pipefish,
Syngnathus typhle
, whose males provide extensive care (nourishment, osmoregulation and oxygenation) to their young in a brood pouch on their bodies. We took advantage of this species' pronounced variation in egg size, correlating positively with female size, and tested the effect of hypoxia (40% dissolved oxygen) versus fully oxygenated (100%) water on embryo size and survival of large versus small eggs after 18 days of paternal brooding. Egg size did not affect embryo survival, regardless of O
2
treatment. While hypoxia affected embryo size negatively, both large and small eggs showed similar reductions in growth. Males in hypoxia ventilated more and males with large eggs swam more, but neither treatment affected their position in the water column. Overall, our results call into question the most common explanation for constrained egg size evolution in aquatic environments.
Competitors as accomplices: seaweed competitors hide corals from predatory sea stars
Authors: Clements Cody S. , Hay Mark E.
Published: August 2015

Indirect biotic effects arising from multispecies interactions can alter the structure and function of ecological communities—often in surprising ways that can vary in direction and magnitude. On Pacific coral reefs, predation by the crown-of-thorns sea star,
Acanthaster planci
, is associated with broad-scale losses of coral cover and increases of macroalgal cover. Macroalgal blooms increase coral–macroalgal competition and can generate further coral decline. However, using a combination of manipulative field experiments and observations, we demonstrate that macroalgae, such as
Sargassum polycystum
, produce associational refuges for corals and dramatically reduce their consumption by
Acanthaster
. Thus, as
Acanthaster
densities increase, macroalgae can become coral mutualists, despite being competitors that significantly suppress coral growth. Field feeding experiments revealed that the protective effects of macroalgae were strong enough to cause
Acanthaster
to consume low-preference corals instead of high-preference corals surrounded by macroalgae. This highlights the context-dependent nature of coral–algal interactions when consumers are common. Macroalgal creation of associational refuges from
Acanthaster
predation may have important implications for the structure, function and resilience of reef communities subject to an increasing number of biotic disturbances.
Resolving the life cycle alters expected impacts of climate change
Authors: Levy Ofir , Buckley Lauren B. , Keitt Timothy H. , Smith Colton D. , Boateng Kwasi O. , Kumar Davina S. , Angilletta Michael J.
Published: August 2015

Recent models predict contrasting impacts of climate change on tropical and temperate species, but these models ignore how environmental stress and organismal tolerance change during the life cycle. For example, geographical ranges and extinction risks have been inferred from thermal constraints on activity during the adult stage. Yet, most animals pass through a sessile embryonic stage before reaching adulthood, making them more susceptible to warming climates than current models would suggest. By projecting microclimates at high spatio-temporal resolution and measuring thermal tolerances of embryos, we developed a life cycle model of population dynamics for North American lizards. Our analyses show that previous models dramatically underestimate the demographic impacts of climate change. A predicted loss of fitness in 2% of the USA by 2100 became 35% when considering embryonic performance in response to hourly fluctuations in soil temperature. Most lethal events would have been overlooked if we had ignored thermal stress during embryonic development or had averaged temperatures over time. Therefore, accurate forecasts require detailed knowledge of environmental conditions and thermal tolerances throughout the life cycle.
Spreading of healthy mood in adolescent social networks
Authors: Hill E. M. , Griffiths F. E. , House T.
Published: August 2015

Depression is a major public health concern worldwide. There is evidence that social support and befriending influence mental health, and an improved understanding of the social processes that drive depression has the potential to bring significant public health benefits. We investigate transmission of mood on a social network of adolescents, allowing flexibility in our model by making no prior assumption as to whether it is low mood or healthy mood that spreads. Here, we show that while depression does not spread, healthy mood among friends is associated with significantly reduced risk of developing and increased chance of recovering from depression. We found that this spreading of healthy mood can be captured using a non-linear complex contagion model. Having sufficient friends with healthy mood can halve the probability of developing, or double the probability of recovering from, depression over a 6–12-month period on an adolescent social network. Our results suggest that promotion of friendship between adolescents can reduce both incidence and prevalence of depression.
Self-discrimination in the tendrils of the vine
Cayratia japonica
is mediated by physiological connection
Authors: Fukano Yuya , Yamawo Akira
Published: August 2015

Although self-discrimination has been well documented, especially in animals, self-discrimination in plants has been identified in only a few cases, such as self-incompatibility in flowers and root discrimination. Here, we report a new form of self-discrimination in plants: discrimination by vine tendrils. We found that tendrils of the perennial vine
Cayratia japonica
were more likely to coil around neighbouring non-self plants than neighbouring self plants in both experimental and natural settings. The higher level of coiling around a physiologically severed self plant compared with that around a physiologically connected self plant suggested that self-discrimination was mediated by physiological coordination between the tendril and the touched plant as reported for self-discrimination in roots. The results highlight the importance of self-discrimination for plant competition not only underground, but also above-ground.
Sickness behaviour associated with non-lethal infections in wild primates
Authors: Ghai Ria R. , Fugère Vincent , Chapman Colin A. , Goldberg Tony L. , Davies T. Jonathan
Published: August 2015

Non-lethal parasite infections are common in wildlife, but there is little information on their clinical consequences. Here, we pair infection data from a ubiquitous soil-transmitted helminth, the whipworm (genus
Trichuris
), with activity data from a habituated group of wild red colobus monkeys (
Procolobus rufomitratus tephrosceles
) in Kibale National Park, Uganda. We use mixed-effect models to examine the relationship between non-lethal parasitism and red colobus behaviour. Our results indicate that red colobus increased resting and decreased more energetically costly behaviours when shedding whipworm eggs in faeces. Temporal patterns of behaviour also changed, with individuals switching behaviour less frequently when whipworm-positive. Feeding frequency did not differ, but red colobus consumption of bark and two plant species from the genus
Albizia
, which are used locally in traditional medicines, significantly increased when animals were shedding whipworm eggs. These results suggest self-medicative plant use, although additional work is needed to verify this conclusion. Our results indicate sickness behaviours, which are considered an adaptive response by hosts during infection. Induction of sickness behaviour in turn suggests that these primates are clinically sensitive to non-lethal parasite infections.
Interaction of influenza virus polymerase with viral RNA in the ‘corkscrew’ conformation
Authors: Flick Ramon , Hobom Gerd
Published: August 2015

The influenza virus RNA (vRNA) promoter structure is known to consist of the 5′- and 3′-terminal sequences of the RNA, within very narrow boundaries of 16 and 15 nucleotides, respectively. A complete set of single nucleotide substitutions led to the previously proposed model of a binary hooked or ‘corkscrew’ conformation for the vRNA promoter when it interacts with the viral polymerase. This functional structure is confirmed here with a complete set of complementary double substitutions, of both the regular A:U and G:C type and also the G:U type of base-pair exchanges. The proposed structure consists of a six base-pair RNA rod in the distal element in conjunction with two stem–loop structures of two short-range base- pairs (positions 2–9; 3–8). These support an exposed tetranucleotide loop within each branch of the proximal element, in an overall oblique organization due to a central unpaired A residue at position 10 in the 5′ sequence. Long-range base-pairing between the entire 5′ and 3′ branches, as required for an unmodified ‘panhandle’ model, has been excluded for the proximal element, while it is known to represent the mode of interaction within the distal element. A large number of short-range base-pair exchanges in the proximal element constitute promoter-up mutations, which show activities several times above that of the wild- type in reporter gene assays. The unique overall conformation and rather few invariant nucleotides appear to be the core elements in vRNA recognition by polymerase and also in viral ribonucleoprotein packaging, to allow discrimination against the background of other RNA molecules in the cell.
Involvement of β2-microglobulin and integrin αvβ3 molecules in the coxsackievirus A9 infectious cycle
Authors: Triantafilou Martha , Triantafilou Kathy , Wilson Keith M. , Takada Yoshikazu , Fernandez Nelson , Stanway Glyn
Published: August 2015

It is becoming apparent that many viruses employ more than one cell surface molecule for their attachment and cell entry. In this study, we have tested the role of integrin αvβ3 and MHC class I molecules in the coxsackievirus A9 (CAV-9) infectious cycle. Binding experiments utilizing CHO cells transfected and expressing human integrin αvβ3, revealed that CAV- 9 particles were able to bind to cells, but did not initiate a productive cell infection. Antibodies specific for integrin αvβ3 molecules significantly reduced CAV-9 infection in susceptible cell lines. Moreover, MAbs specific for β2- microglobulin (β2-m) and MHC class I molecules completely inhibited CAV-9 infection. To assess the effect of these antibodies on virus binding, we analysed CAV-9 binding by flow cytometry in the presence of β2-m- or integrin αvβ3-specific antibodies. The results showed a reduction in CAV-9 binding in the presence of integrin αvβ3- specific antibodies while there was no reduction in the presence of β2-m-specific MAb. Taken together, these data suggest that integrin αvβ3 is required for CAV-9 attachment but is not sufficient for cell entry, while β2-m, although not directly involved in CAV-9 binding, plays a post- attachment role in the CAV-9 infectious process, possibly being involved in virus entry.
Interaction of YB-1 with human immunodeficiency virus type 1 Tat and TAR RNA modulates viral promoter activity
Authors: Ansari Sameer A. , Safak Mahmut , Gallia Gary L. , Sawaya Bassel E. , Amini Shohreh , Khalili Kamel
Published: August 2015

Transcriptional regulation of the human immunodeficiency virus type 1 (HIV-1) genome is mediated by viral and cellular factors. TAR, an unusual RNA regulatory element with a stem–bulge–loop structure at the 5′ ends of all nascent viral transcripts is critical for HIV-1 transcription. TAR is the target for Tat, a viral transcription factor encoded early in the HIV-1 life-cycle and essential for gene expression. Evidence demonstrating the interaction of a cellular ssDNA/RNA binding protein, YB-1, with TAR through a region which is important for Tat interaction is presented. Interestingly, results from protein–protein interaction studies revealed that YB-1 can also form a complex with Tat. Results from mapping experiments suggest that while the region spanning aa 125–203 within YB-1 is essential for its association with TAR, a truncated YB-1 spanning aa 1–125 can weakly bind to Tat. Functionally, overexpression of full-length YB-1 enhanced Tat-induced activation of the HIV-1 minimal promoter containing TAR sequences, whereas mutant YB-1 with no ability to bind to Tat and TAR failed to affect Tat-mediated activation. Expression of mutant YB-1(1–125), which binds to Tat but not RNA, decreased Tat- mediated enhancement of virus transcription. These observations suggest that while full-length YB-1 may function as a facilitator and, by interaction with both Tat and TAR, increase the level of Tat:TAR association, mutant YB-1 with no TAR binding activity, by complexing with Tat, may prevent Tat interaction with TAR. The importance of these findings in light of the proposed mechanism of Tat function is discussed.
The U28 ORF of human herpesvirus-7 does not encode a functional ribonucleotide reductase R1 subunit
Authors: Sun Yunming , Conner Joe
Published: August 2015

Herpesvirus ribonucleotide reductases, essential for the de novo synthesis of viral DNA, are composed of two non-identical subunits, termed R1 and R2. The U28 ORF from human herpesvirus-7 has been classified, by sequence comparisons, as a homologue of the R1 subunit from ribonucleotide reductase but no R2 ORF is present. Detailed analysis of the U28 amino acid sequence indicated that a number of essential R1 catalytic residues are absent. Cloning and expression of the U28 protein in E. coli and its subsequent characterization in subunit interaction and enzyme activity assays confirmed that it is not a functional equivalent of a herpesvirus R1. In the absence of the R2 gene, we propose that the R1 ORF has evolved a distinct, as yet unidentified, function not only in human herpesvirus-7 but also in other human betaherpesviruses.
The murine gammaherpesvirus-68 M11 protein inhibits Fas- and TNF- induced apoptosis
Authors: Wang Guang-Hua , Garvey Tara L. , Cohen Jeffrey I.
Published: August 2015

The murine gammaherpesvirus-68 (MHV-68) M11 gene encodes a protein predicted to have limited homology to the bcl-2 family of proteins. Unlike most of the other viral bcl-2 homologues, which have both BH1 and BH2 domains conserved with respect to bcl-2, the M11 protein has a BH1 domain, but apparently lacks a BH2 domain. Transfection of HeLa cells with an epitope-tagged MHV-68 M11 construct showed that the protein is predominantly located in the cytoplasm of cells. In HeLa cells, M11 inhibited apoptosis induced by anti-Fas antibody and by TNF-α. Thus, despite its limited conservation with respect to other bcl-2 family members, the MHV-68 M11 protein is a potent inhibitor of apoptosis.
Characterization of the Epstein–Barr virus BALF2 promoter
Authors: Hung Chien-Hui , Liu Shih-Tung
Published: August 2015

BALF2, which encodes the major DNA-binding protein of Epstein–Barr virus (EBV), is expressed during the early stage of the lytic cycle. The location of the BALF2 promoter was identified by primer extension, which indicated that the transcription start is located at nucleotide 164,782 of the EBV genome. Transfection analyses revealed that, similar to other EBV early promoters, the BALF2 promoter is activated by the EBV-encoded transcription factors Rta and Zta. The promoter is also synergistically activated if both transcription factors are present in B lymphocytes and in epithelial cells. Deletion analysis and electrophoretic mobility-shift assay revealed that the region between nucleotides −134 and −64 contains Zta- response elements and the region between nucleotides −287 and −254 contains Rta-response elements. This study demonstrates the importance of Rta and Zta in regulating the transcription of EBV early genes.
Oral transmission and early lymphoid tropism of chronic wasting disease PrPres in mule deer fawns (Odocoileus hemionus )
Authors: Sigurdson Christina J. , Williams Elizabeth S. , Miller Michael W. , Spraker Terry R. , O’Rourke Katherine I. , Hoover Edward A.
Published: August 2015

Mule deer fawns (Odocoileus hemionus) were inoculated orally with a brain homogenate prepared from mule deer with naturally occurring chronic wasting disease (CWD), a prion-induced transmissible spongiform encephalopathy. Fawns were necropsied and examined for PrP res, the abnormal prion protein isoform, at 10, 42, 53, 77, 78 and 80 days post-inoculation (p.i.) using an immunohistochemistry assay modified to enhance sensitivity. PrPres was detected in alimentary-tract-associated lymphoid tissues (one or more of the following: retropharyngeal lymph node, tonsil, Peyer’s patch and ileocaecal lymph node) as early as 42 days p.i. and in all fawns examined thereafter (53 to 80 days p.i.). No PrPres staining was detected in lymphoid tissue of three control fawns receiving a control brain inoculum, nor was PrPres detectable in neural tissue of any fawn. PrPres-specific staining was markedly enhanced by sequential tissue treatment with formic acid, proteinase K and hydrated autoclaving prior to immunohistochemical staining with monoclonal antibody F89/160.1.5. These results indicate that CWD PrP res can be detected in lymphoid tissues draining the alimentary tract within a few weeks after oral exposure to infectious prions and may reflect the initial pathway of CWD infection in deer. The rapid infection of deer fawns following exposure by the most plausible natural route is consistent with the efficient horizontal transmission of CWD in nature and enables accelerated studies of transmission and pathogenesis in the native species.
PrP genotypes of captive and free-ranging Rocky Mountain elk (Cervus elaphus nelsoni) with chronic wasting disease
Authors: O’Rourke K. I. , Besser T. E. , Miller M. W. , Cline T. F. , Spraker T. R. , Jenny A. L. , Wild M. A. , Zebarth G. L. , Williams E. S.
Published: August 2015

The PrP gene encodes the putative causative agent of the transmissible spongiform encephalopathies (TSEs), a heterogeneous group of fatal, neurodegenerative disorders including human Creutzfeldt–Jakob disease, bovine spongiform encephalopathy, ovine scrapie and chronic wasting disease (CWD) of North American deer and elk. Polymorphisms in the PrP gene are associated with variations in relative susceptibility, pathological lesion patterns, incubation times and clinical course of TSEs of humans, mice and sheep. Sequence analysis of the PrP gene from Rocky Mountain elk showed only one amino acid change (Met to Leu at cervid codon 132). Homozygosity for Met at the corresponding polymorphic site (Met to Val) in humans (human codon 129) predisposes exposed individuals to some forms of Creutzfeldt–Jakob disease. In this study, Rocky Mountain elk homozygous for PrP codon 132 Met were over-represented in both free- ranging and farm-raised CWD-affected elk when compared to unaffected control groups.
The nucleopolyhedroviruses of Rachiplusia ou and Anagrapha falcifera are isolates of the same virus
Authors: Harrison Robert L. , Bonning Bryony C.
Published: August 2015

The 7·8 kb EcoRI-G fragment of Rachiplusia ou multicapsid nucleopolyhedrovirus (RoMNPV), containing the polyhedrin gene, was cloned and sequenced. The sequence of the fragment was 92·3% identical to the sequence of the corresponding region in the Autographa californica (Ac)MNPV genome. A comparison of the EcoRI-G sequence with other MNPV sequences revealed that RoMNPV was most closely related to AcMNPV. However, the predicted amino acid sequence of RoMNPV polyhedrin shared more sequence identity with the polyhedrin of Orygia pseudotsugata MNPV. In addition, the RoMNPV sequence was almost completely identical (99·9%) to a previously published 6·3 kb sequence of Anagrapha falcifera MNPV (AfMNPV). The Eco RI and HindIII restriction fragment profiles of RoMNPV and AfMNPV also were nearly identical, with an additional EcoRI band detected in RoMNPV DNA. Bioassays of these viruses with three different hosts (the European corn borer, Ostrinia nubilalis H übner, the corn earworm, Helicoverpa zea Boddie, and the tobacco budworm, Heliothis virescens Fabricius) failed to detect any differences in the biological activities of RoMNPV and AfMNPV. These results indicate that RoMNPV and AfMNPV are different isolates of the same virus. The taxonomic relationship of Ro/AfMNPV and AcMNPV is discussed.
Transformation of tobacco and potato with cDNA encoding the full-length genome of Potato leafroll virus: evidence for a novel virus distribution and host effects on virus multiplication
Authors: Franco-Lara Liliana F. , McGeachy Kara D. , Commandeur Uli , Martin Robert R. , Mayo Mike A. , Barker Hugh
Published: August 2015

A full-length cDNA copy of the genome of Potato leafroll virus (PLRV) was introduced into the genome of tobacco and potato plants by Agrobacterium tumefaciens-mediated transformation. Transgenic lines were obtained in which the transgene was readily detected by PCR with DNA extracted from T1 tobacco seedlings and clonally multiplied potato plants. PLRV-specific genomic and sub- genomic RNAs, coat protein antigen and virus particles were detected in transgenic plants. Aphids fed on the transgenic tobacco plants readily transmitted PLRV to test plants. Infected transgenic tobacco plants, like non-transgenic (WT) PLRV-infected plants, displayed no symptoms of the infection but transgenic plants of potato were severely stunted. In parallel tests, the mean PLRV titres in WT tobacco plants and transgenic tobacco plants were 600 and 630 ng virus/g leaf, respectively, although differences in PLRV titres among transgenic plants were much greater than those among infected WT plants. In similar tests with potato, the mean PLRV titre of WT plants was 50 ng virus/g leaf whereas higher concentrations (up to 3400 ng virus/g leaf) accumulated in transgenic potato plants. In tissue prints of stems, PLRV was detected in similar proportions of phloem cells in transgenic and infected WT plants. In transgenic tobacco and potato plants, but not in infected WT plants, a few stem epidermal cells also contained virus. From tissue prints of transgenic tobacco leaves, it was estimated that about one in 40000 mesophyll cells contained virus, but in transgenic potato, a greater proportion of mesophyll cells was infected.
Expression of bovine viral diarrhoea virus glycoprotein E2 by bovine herpesvirus-1 from a synthetic ORF and incorporation of E2 into recombinant virions
Authors: Schmitt Jutta , Becher Paul , Thiel Heinz-Jürgen , Keil Günther M.
Published: August 2015

Expression cassettes containing the codons for the pestivirus E rns signal peptide (Sig) followed by a chemically synthesized ORF that encoded the bovine viral diarrhoea virus (BVDV) strain C86 glycoprotein E2, a class I membrane glycoprotein, were constructed with and without a chimeric intron sequence immediately upstream of the translation start codon, and incorporated into the genome of bovine herpesvirus-1 (BHV-1). The resulting recombinants, BHV- 1/SigE2syn and BHV-1/SigE2syn-intron, expressed comparable quantities of glycoprotein E2, and Northern blot hybridizations indicated that the presence of the intron did not increase significantly the steady-state levels of transcripts encompassing the SigE2syn ORF. In BHV-1/SigE2syn- infected cells, the 54 kDa E2 glycoprotein formed a dimer with an apparent molecular mass of 94 kDa, which was further modified to a 101 kDa form found in the envelope of recombinant virus particles. Penetration kinetics and single-step growth curves indicated that the incorporation of the BVDV E2 glycoprotein in the BHV-1 envelope, which apparently did not require BHV-1-specific signals, interfered with entry into target cells and egress of progeny virions. These results demonstrate that a pestivirus glycoprotein can be expressed efficiently by BHV-1 and incorporated into the viral envelope. BHV-1 thus represents a promising tool for the development of efficacious live and inactivated BHV-1-based vector vaccines.
Free thiol groups are essential for infectivity of human cytomegalovirus
Authors: Mirazimi Ali , Mousavi-Jazi Mehrdad , Sundqvist Vivi-Anne , Svensson Lennart
Published: August 2015

The membrane-impermeable thiol blocker 5′5-dithiobis 2- nitrobenzoic acid (DTNB) blocked infectivity of human cytomegalovirus (CMV) although the virus still bound to cells. DTNB-treated CMV regained 65% of its infectivity after incubation with the disulfide bond-reducing agent dithiothreitol. These observations suggest that free thiol groups on CMV are required for infectivity and may participate in disulfide bond formation during virus entry.
Forced recombination of Ψ-modified murine leukaemia virus-based vectors with murine leukaemia-like and VL30 murine endogenous retroviruses
Authors: Mikkelsen Jacob Giehm , Lund Anders H. , Duch Mogens , Pedersen Finn Skou
Published: August 2015

Co-encapsidation of retroviral RNAs into virus particles allows for the generation of recombinant proviruses through events of template switching during reverse transcription. By use of a forced recombination system based on recombinational rescue of replication- defective primer binding site-impaired Akv–MLV-derived vectors, we here examine putative genetic interactions between vector RNAs and copackaged endogenous retroviral RNAs of the murine leukaemia virus (MLV) and VL30 retroelement families. We show (i) that MLV recombination is not blocked by nonhomology within the 5′ untranslated region harbouring the supposed RNA dimer-formingcis-elements and (ii) that copackaged retroviral RNAs can recombine despite pronounced sequence dissimilarity at the cross-over site(s) and within parts of the genome involved in RNA dimerization, encapsidation and strand transferring during reverse transcription. We note that recombination-based rescue of primer binding site knock-out retroviral vectors may constitute a sensitive assay to register putative genetic interactions involving endogenous retroviral RNAs present in cells of various species.
An analysis of the role of neuraminidase in the receptor-binding activity of influenza B virus: the inhibitory effect of Zanamivir on haemadsorption
Authors: Luo C. , Nobusawa E. , Nakajima K.
Published: August 2015

We analysed the role of neuraminidase (NA) on haemadsorption by the haemagglutinin (HA) protein of influenza B virus. The influenza B virus mutant ts-7 has a temperature-sensitive mutation in the NA protein. At high temperature, cells infected with this virus did not exhibit haemadsorption activity, but the addition of bacterial neuraminidase (bNA) restored haemadsorption activity. COS cells transfected with HA cDNAs of B/Kanagawa/73 or B/Lee/40 virus showed no evidence of haemadsorption. However, with the addition of bNA or co- transfection with NA cDNA of the B/Lee/40 virus, haemadsorption was observed. Experiments with point-mutated HA cDNAs of B/Lee/40 virus showed that two N-acetyl glycosylation sites at amino acid residues 160 and 217 were responsible for the inability of the HA protein to adsorb to erythrocytes. These results indicated that haemadsorption by the HA protein of influenza B virus required the involvement of NA. Because the NA inhibitor Zanamivir was reported not to penetrate cells, we investigated the action of this inhibitor and found that Zanamivir inhibited haemadsorption on MDCK cells infected with B/Kanagawa/73 or B/Lee/40 virus. After removing Zanamivir by washing, the addition of bNA restored the haemadsorption activity on the infected cells. Scanning electron microscopy indicated that at 0·4 μM Zanamivir, HA protein did not adsorb to erythrocytes but retained the ability to aggregate virions. However, at 4 μM Zanamivir, distinct virion formation could not be observed.
Transmission of the 263K scrapie strain by the dental route
Authors: lngrosso Loredana , Pisani Flavio , Pocchiari Maurizio
Published: August 2015

Apart from a few cases of iatrogenic and familial human transmissible spongiform encephalopathies (TSEs) or prion diseases, the cause of Creutzfeldt–Jakob disease (CJD) remains unknown. In this paper we investigated the possibility that dental procedures may represent a potential route of infection. This was assessed by using the experimental model of scrapie in hamster. In the first part of this study we found that after intraperitoneal inoculation, oral tissues commonly involved in dental procedures (gingival and pulp tissues) bore a substantial level of infectivity. We also found high scrapie infectivity in the trigeminal ganglia, suggesting that the scrapie agent had reached the oral tissues through the sensitive terminal endings of the trigeminal nerves. In the second part of the study we inoculated a group of hamsters in the tooth pulp and showed that all of them developed scrapie disease. In these animals, we detected both infectivity and the pathological prion protein (PrPsc) in the trigeminal ganglion homolateral to the site of injection but not in the controlateral one. This finding suggests that the scrapie agent, and likely other TSE agents as well, spreads from the buccal tissues to the central nervous system through trigeminal nerves. Although these findings may not apply to humans affected by TSEs, they do raise concerns about the possible risk of transmitting these disorders through dental procedures. Particular consideration should be taken in regard to new variant CJD patients because they may harbour more infectivity in peripheral tissues than sporadic CJD patients.
First seroepidemiological study and phylogenetic characterization of human T-cell lymphotropic virus type I and II infection among Amerindians in French Guiana
Authors: Talarmin Antoine , Vion Bruno , Ureta-Vidal Abel , Du Fou Guénola , Marty Christian , Kazanji Mirdad
Published: August 2015

We investigated the serological, epidemiological and molecular aspects of human T-cell lymphotropic virus type I and II (HTLV-I/II) infection in the Amerindian populations of French Guiana by testing 847 sera. No HTLV-II antibodies were detected, but five individuals (0·59%) were seropositive for HTLV-I. Analysis of the nucleotide sequences of 522 bp of the env gene and the compete LTR showed that all of the strains from French Guiana belonged to the cosmopolitan subtype A. The similarities were greater between Amerindian and Creole strains than between Amerindian and Noir-Marron strains or than between Creole and Noir-Marron strains. Phylogenetic analysis showed two clusters: one of strains from Amerindians and Creoles, which belong to the transcontinental subgroup, and the other of strains from Noirs-Marrons, belonging to the West African subgroup. Our results suggest that the Amerindian HTLV-I strains are of African origin.
Mapping the binding domains on decay accelerating factor (DAF) for haemagglutinating enteroviruses: implications for the evolution of a DAF-binding phenotype
Authors: Powell Robert M. , Ward Trevor , Goodfellow Ian , Almond Jeffrey W. , Evans David J.
Published: August 2015

Decay accelerating factor (DAF) functions as a cell attachment receptor for a wide range of human enteroviruses, the interaction accounting for the haemagglutination phenotype exhibited by many members of this family. Haemagglutination inhibition assays using purified truncated soluble DAF (sDAF) receptors and short consensus repeat (SCR) domain-specific antibodies have been used to determine the domain(s) of DAF to which the viruses bind. Further sDAF-mediated virus neutralization and biosensor analysis have been used to confirm the virus-binding domains of DAF. Of the four distinct clusters of human enteroviruses, three contain representatives that bind DAF. The majority of DAF-binding enteroviruses occupy the ‘CBV-like’ cluster, and require SCR domains 2–4 for DAF binding. In contrast, the DAF-binding representatives of the ‘ENV70-like’ and ‘PV-like’ clusters require SCR1 for DAF interaction. These studies confirm that DAF binding is a widespread characteristic amongst phylogenetically divergent clusters within the enteroviruses and suggest that the ability to bind DAF may have evolved more than once within this group of viruses.
Molecular characterization of human group C rotavirus genes 6, 7 and 9
Authors: James Vivienne L. A. , Lambden Paul R. , Deng Yu , Caul E. Owen , Clarke Ian N.
Published: August 2015

Genes 6, 7 and 9 of human group C rotavirus ‘Bristol’ strain, encoding non-structural proteins (NSP) 3, 1 and 2, respectively, were cloned and sequenced. Human group C rotavirus genome segment 6 is 1350 bp and contains a single ORF of 1231 nucleotides (encoding 402 amino acids). Genome segment 7 is 1270 bp and encodes a protein of 394 amino acids and genome segment 9 is 1037 bp and encodes a 312 amino acid protein. The human group C rotavirus genes 6, 7 and 9 showed 78, 67 and 88% sequence identity, respectively, to the corresponding porcine group C rotavirus genes. The derived protein sequences were compared with those of the porcine ‘Cowden’ group C and mammalian group A rotavirus strains. The human group C rotavirus NSP1 protein sequence is one amino acid longer than the porcine group C equivalent. In common with group A and porcine group C rotaviruses, the human group C rotavirus NSP1 protein has a zinc finger motif. Human group C rotavirus NSP2 has two hydrophobic heptad repeat regions, a basic, RNA-binding domain and a basic, proline-rich region. Human group C rotavirus NSP3 has both single- and double-stranded RNA-binding domains and several hydrophobic heptad repeat regions, one of which forms a leucine zipper. This work completes the molecular characterization of the non-structural proteins of a human group C rotavirus. Phylogenetic analysis of all the non-structural genes of group A, B and C rotaviruses suggests that these viruses have diverged at a constant rate from a common ancestor.
Characterization of the DNA polymerase loci of the novel porcine lymphotropic herpesviruses 1 and 2 in domestic and feral pigs
Authors: Ulrich Sven , Goltz Michael , Ehlers Bernhard
Published: August 2015

Two novel porcine gammaherpesviruses, porcine lymphotropic herpesviruses 1 and 2 (PLHV-1 and -2), have been detected by amplification of short DNA polymerase (DPOL) sequences from blood and spleen of domestic pigs while searching for unknown herpesviruses in pigs as possible risk factors in xenotransplantation. In the present study, the DPOL genes of the two viruses and the open reading frames (ORFs) that follow in the downstream direction were amplified by PCR-based genome walking from adaptor-ligated restriction fragment libraries of porcine spleen samples. The sequences determined for the two PLHVs exhibited a very low G+C content (37 mol%) and a marked suppression of the CpG dinucleotide frequency. The DPOL proteins encoded were 95% identical and showed a close relationship (60% identity) to the DPOL protein of a ruminant gammaherpesvirus, alcelaphine herpesvirus 1 (AlHV-1). This was confirmed by phylogenetic analyses of the conserved regions of the two PLHV DPOL proteins. The PLHV ORFs downstream of DPOL exhibited 83% identity to each other and ≫50% similarity to ORF A5, the position equivalent of AlHV-1. From these data, the PLHVs can be firmly classified to the subfamily Gammaherpesvirinae. To find a natural reservoir for the PLHVs, organs of feral pigs were screened with five different PCR assays, targetting either the DPOL gene or 3′-flanking sequences. In all samples, PLHV sequences were detected that originated predominantly from PLHV-2, suggesting the possibility of virus transfer between feral and domestic pig populations.
Identification of a new promoter in the early region of the human papillomavirus type 16 genome
Authors: Braunstein T. H. , Madsen B. S. , Gavnholt B. , Rosenstierne M. W. , Koefoed Johnsen C. , Norrild B.
Published: August 2015

Transcription of the human papillomavirus type 16 (HPV-16) genome is controlled by several promoters; the P97 promoter is considered to be the main one. An additional promoter has been identified within the E7 ORF as well as an antisense promoter just upstream of the L2 ORF. The significance of these promoters for early and late gene expression and their activity related to cell differentiation is not known in detail. Identification of two new, previously undescribed transcription start sites at nt 542 just upstream of the E7 ORF and at nt 611 within the E7 ORF is reported. The promoter responsible for the start site at nt 542 (P542) was active in SiHa, HeLa and C33A cells. Very low promoter activity was found upstream of the nt 611 start site. The E7 protein has previously been shown to be synthesized from a polycistronic mRNA encoding both the E6 and E7 proteins under the control of the P97 promoter. The data reported in the present paper suggest that promoter P542 may control synthesis of the E7 oncoprotein from a monocistronic mRNA.
Induction of apoptosis after switch-on of the hepatitis B virus X gene mediated by the Cre/loxP recombination system
Authors: Shintani Yoshizumi , Yotsuyanagi Hiroshi , Moriya Kyoji , Fujie Hajime , Tsutsumi Takeya , Kanegae Yumi , Kimura Satoshi , Saito Izumu , Koike Kazuhiko
Published: August 2015

The HBx protein of hepatitis B virus is a multifunctional protein that is implicated in the pathogenesis of hepatocellular carcinoma by regulating gene transcription, causing cell proliferation and, as shown recently, inducing cell death. However, analysis of the effects of HBx in stable cultured cell clones has been hampered because only cell lines that adapted to the effects of HBx were selected during the establishment of cell clones. Here, we describe a system in which transcription of the X gene of hepatitis B virus is switched on by the use of the site-specific Cre recombinase. Two human liver cell lines, HLF and HepG2, were used, the former with a mutant p53 allele and the latter with wild-type p53. The stable cell clones isolated, which carried the X gene in a transcriptionally silent state, were infected with recombinant adenovirus carrying Cre recombinase. Ninety-six hours after adenovirus infection, cell clones that expressed HBx had undergone TUNEL-positive cell death with characteristics of apoptosis. Apoptosis was induced despite concomitant inactivation of the p53 protein as a result of its cytoplasmic translocation by HBx. In contrast, neither the X gene-carrying cells infected with wild-type adenovirus nor various control cells infected with Cre-expressing adenovirus exhibited apoptosis. These results indicate that the expression of HBx protein leads to liver cell apoptosis independently of the p53 pathway. The significance of HBx-induced apoptosis in natural infection is unclear, but it may contribute to the development of hepatitis and serve to spread progeny virus to neighbouring cells while evading the host immune responses.
Context of the coat protein DAG motif affects potyvirus transmissibility by aphids
Authors: López-Moya J. J. , Wang R. Y. , Pirone T. P.
Published: August 2015

Previous work with tobacco vein mottling virus (TVMV) has established that a highly conserved three amino acid motif, asp-ala-gly (DAG), located near the N terminus of the coat protein (CP), is important for aphid transmission. However, several other potyviruses which have motifs other than DAG are aphid-transmissible. Creation of these motifs in TVMV through site-directed mutagenesis failed to render TVMV aphid-transmissible from infected plants, and the creation of a putative complementary motif in the helper component did not restore transmissibility. In an isolate of tobacco etch virus (TEV) that contains two consecutive DAG motifs separated by a single ala, transmissibility was abolished or reduced by mutations affecting the first motif, whereas mutations in the second motif had little or no effect. In a TEV mutant made non-transmissible due to an altered first motif, substitution of val for ala in the position immediately before the second DAG restored transmissibility, whereas changing val to ala in the location prior to the first DAG resulted in reduced TEV transmissibility. In contrast, a val to ala change in the position preceding the single DAG motif of TVMV did not affect transmission. Creation of another DAG motif at the beginning of the TVMV CP core, in a position where certain other potyviruses have a second DAG motif, did not restore transmissibility. Our results suggest that the mere presence of a DAG motif does not guarantee transmissibility and that the context in which the DAG or equivalent motif is found plays a role in the process.
Sequence and organization of the Spodoptera exigua multicapsid nucleopolyhedrovirus genome
Authors: IJkel Wilfred F. J. , van Strien Elisabeth A. , Heldens Jacobus G. M. , Broer René , Zuidema Douwe , Goldbach Rob W. , Vlak Just M.
Published: August 2015

The nucleotide sequence of the DNA genome of Spodoptera exigua multicapsid nucleopolyhedrovirus (SeMNPV), a group II NPV, was determined and analysed. The genome contains 135611 bp and has a G+C content of 44 mol%. Computer-assisted analysis revealed 139 ORFs of 150 nucleotides or larger; 103 have homologues in Autographa californica MNPV (AcMNPV) and a further 16 have homologues in other baculoviruses. Twenty ORFs are unique to SeMNPV. Major differences in SeMNPV gene content and arrangement were found compared with the group I NPVs AcMNPV, Bombyx mori (Bm) NPV and Orgyia pseudotsugata (Op) MNPV and the group II NPV Lymantria dispar (Ld) MNPV. Eighty-five ORFs were conserved among all five baculoviruses and are considered as candidate core baculovirus genes. Two putative p26 and odv-e66 homologues were identified in SeMNPV, each of which appeared to have been acquired independently and not by gene duplication. The SeMNPV genome lacks homologues of the major budded virus glycoprotein gene gp64, the immediate-early transactivator ie-2 and bro (baculovirus repeat ORF) genes that are found in AcMNPV, BmNPV, OpMNPV and LdMNPV. Gene parity analysis of baculovirus genomes suggests that SeMNPV and LdMNPV have a recent common ancestor and that they are more distantly related to the group I baculoviruses AcMNPV, BmNPV and OpMNPV. The orientation of the SeMNPV genome is reversed compared with the genomes of AcMNPV, BmNPV, OpMNPV and LdMNPV. However, the gene order in the ‘central’ part of baculovirus genomes is highly conserved and appears to be a key feature in the alignment of baculovirus genomes.
Bacteriophage-like particles associated with the gene transfer agent of Methanococcus voltae PS
Authors: Eiserling F. , Pushkin A. , Gingery M. , Bertani G.
Published: August 2015

The methanogenic archaeobacterium Methanococcus voltae (strain PS) is known to produce a filterable, DNase-resistant agent (called VTA, for voltae transfer agent), which carries very small fragments (4400 bp) of bacterial DNA and is able to transduce bacterial genes between derivatives of the strain. Examination by electron microscopy of two preparations of VTA that were concentrated and partially purified by different methods showed virus-like particles with isometric heads, about 40 nm in diameter, and with 61 nm long tails. These particles co-sedimented with the minute bacteriophage ϕX174 in a sucrose density gradient.
Co-expression of a trans-dominant negative mutant of the human immunodeficiency virus type 1 (HIV-1) Rev protein affects the Rev-dependent splicing pattern and expression of HIV-1 RNAs
Authors: Szilvay Anne Marie , Bøe Stig-Ove , Kalland Karl-Henning
Published: August 2015

Trans-dominant negative mutants of the human immunodeficiency virus type 1 (HIV-1) regulatory protein Rev inhibit the function of wild-type Rev in a dose-dependent manner. This was previously shown to be caused by nuclear retention of the wild-type protein. In the present work, further analysis of thetrans-dominant negative effect was performed using cotransfection experiments with different constructs encoding HIV-1 Rev and viral structural proteins together with a plasmid encoding atrans-dominant negative Rev mutant. Thus, one species of pre-mRNA was transcribed from the reporter plasmids. This pre-mRNA was then either spliced or exported by Rev as unspliced RNA for translation of the HIV structural proteins. An immunofluorescence assay and Western blot analysis were used for analysis of protein expression.In situhybridization was applied for labelling of unspliced mRNA in transfected cells, and RNase protection analysis was used to determine the relative amount of unspliced versus spliced mRNAs. The experiments confirmed that thetrans-dominant negative mutant inhibited nuclear export of unspliced mRNA. It was, in addition, demonstrated for the first time that thetrans-dominant negative mutant also affected a Rev-dependent regulatory step connected with viral pre-mRNA splicing. As a consequence, proteins expressed from unspliced and singly spliced HIV mRNAs decreased while there was an increase in protein products encoded by spliced and alternatively spliced mRNAs.
Manganese cations increase the mutation rate of human immunodeficiency virus type 1 ex vivo
Authors: Vartanian Jean-Pierre , Sala Monica , Henry Michel , Wain-Hobson Simon , Meyerhans Andreas
Published: August 2015

Human immunodeficiency virus (HIV) reverse transcription is an error-prone process with an overall mutation rate of ∼3·4×10−5per base per replication cycle. This rate can be modulated by changes in different components of the retrotranscription reaction. In particular,in vitrosubstitution of magnesium cations (Mg2+) by manganese cations (Mn2+) has been shown to increase misincorporation of deoxynucleotide triphosphates (dNTPs) and to alter substrate specificity. Here, it is shown that Mn2+also increases the HIV mutation rateex vivo. Treatment of permissive cells with Mn2+and subsequent HIV infection resulted in at least 6-fold and 10-fold increases in the mutant and mutation frequencies respectively, thus illustrating a further example of how to influence HIV genetic variation.