In vivo studies of genomic packaging in the dsRNA bacteriophage Φ8

BACKGROUND: Φ8 is a bacteriophage containing a genome of three segments of double-stranded RNA inside a polyhedral capsid enveloped in a lipid-containing membrane. Plus strand RNA binds and is packaged by empty procapsids. Whereas Φ6, another member of the Cystoviridae, shows high stringency, serial dependence and precision in its genomic packaging in vitro and in vivo, Φ8 packaging is more flexible. Unique sequences (pac) near the 5' ends of plus strands are necessary and sufficient for Φ6 genomic packaging and the RNA binding sites are located on P1, the major structural protein of the procapsid. RESULTS: In this paper the boundaries of the Φ8 pac sequences have been explored by testing the in vivo packaging efficacy of transcripts containing deletions or changes in the RNA sequences. The pac sequences have been localized to the 5' untranslated regions of the viral transcripts. Major changes in the pac sequences are either tolerated or ameliorated by suppressor mutations in the RNA sequence. Changes in the genomic packaging program can be established as a result of mutations in P1, the major structural protein of the procapsid and the determinant of RNA binding specificity. CONCLUSION: Although Φ8 is distantly related to bacteriophage Φ6, and does not show sequence similarity, it has a similar genomic packaging program. This program, however, is less stringent than that of Φ6

Characterization of Φ2954, a newly isolated bacteriophage containing three dsRNA genomic segments

Abstract Background Bacteriophage Φ12 is a member of the Cystoviridae and is distinct from Φ6, the first member of that family. We have recently isolated a number of related phages and five showed high similarity to Φ12 in the amino acid sequences of several proteins. Bacteriophage Φ2954 is a member of this group. Results Φ2954 was isolated from radish leaves and was found to have a genome of three segments of double-stranded RNA (dsRNA), placing it in the Cystoviridae. The base sequences for many of the genes and for the segment termini were similar but not identical to those of bacteriophage Φ12. However, the host specificity was for the type IV pili of Pseudomonas syringae HB10Y rather than for the rough LPS to which Φ12 attaches. Reverse genetics techniques enabled the production of infectious phage from cDNA copies of the genome. Phage were constructed with one, two or three genomic segments. Phage were also produced with altered transcriptional regulation. Although the pac sequences of Φ2954 show no similarity to those of Φ12, segment M of Φ2954 could be acquired by Φ12 resulting in a change of host specificity. Conclusions We have isolated a new member of the bacteriophage family Cystoviridae and find that although it shows similarity to other members of the family, it has unique properties that help to elucidate viral strategies for genomic packaging and gene expression.</p

Characterization of Φ8, a Bacteriophage Containing Three Double-Stranded RNA Genomic Segments and Distantly Related to Φ6

AbstractThe three double-stranded RNA genomic segments of bacteriophage Φ8 were copied as cDNA, and their nucleotide sequences were determined. Although the organization of the genome is similar to that of Φ6, there is no similarity in either the nucleotide sequences or the amino acid sequences, with the exception of the motifs characteristic of viral RNA polymerases that are found in the presumptive polymerase sequence. Several features of the viral proteins differ markedly from those of Φ6. Although both phages are covered by a lipid-containing membrane, the protein compositions are very different. The most striking difference is that protein P8, which constitutes a shell around the procapsid in Φ6, is part of the membrane in Φ8. The host attachment protein consists of two peptides rather than one and the phage attaches directly to the lipopolysaccharide of the host rather than to a type IV pilus. The host range of Φ8 includes rough strains of Salmonella typhimurium and of pseudomonad

Effect of nitroglycerin during hemodynamic estimation of valve orifice in patients with mitral stenosis

In patients with mitral stenosis, valve orifice calculations using pulmonary capillary wedge pressure as a substitute for left atrial pressure may overestimate the severity of disease. Previous studies have shown that mitral valve area determined from transseptal left atrial pressure measurements exceeds that area derived from pulmonary wedge pressure measurements. This is probably due to pulmonary venoconstriction, which is reversed by nitroglycerin. Nitroglycerin, 0.4 mg, was administered sublingually to 20 patients with mitral valve disease during preoperative cardiac catheterization using the pulmonary capillary wedge pressure as the proximal hydraulic variable. At the time of peak hypotensive effect, 3 to 5 minutes after nitroglycerin administration, the mean pulmonary capillary wedge pressure decreased from 23 ± 2 (mean ± SEM) to 19 ± 2 mm Hg (p < 0.005). The mean diastolic transmitral pressure gradient (12.6 ± 1.2 mm Hg before and 11.5 ± 1.0 mm Hg after nitroglycerin; p = NS) and cardiac output (4.0 ± 0.3 to 4.1 ± 0.3 liters/min; p = NS) did not change significantly. Nevertheless, the hemodynamic mitral orifice area, calculated using the Gorlin formula, increased from 0.8 ± 0.1 to 1.1 ± 0.2 cm2(p < 0.05). In 12 patients with isolated mitral stenosis, without regurgitation, the mitral valve orifice area after nitroglycerin was 0.4 ± 0.2 cm2larger than it was before drug administration (p < 0.05).Administration of nitroglycerin during evaluation of mitral stenosis eliminates pulmonary venoconstriction, which raises the pulmonary capillary wedge pressure above the left atrial pressure in some patients. Nitroglycerin may add diagnostic accuracy without transseptal catheterization. Whether this response to nitroglycerin has direct therapeutic value in patients with mitral valve obstruction has yet to be determined

Back to the Future: How UK-based news organisations are rediscovering objectivity

The emergence of 'fake news' during the tumultuous Brexit referendum and Trump election campaign has sent news organisations scurrying to set up special teams of journalists to debunk deliberately misleading stories and verify facts. This paper examines the steps being taken to counter the spate of false news stories being spread through social media and asks whether normative values of objectivity are about to enjoy a comeback. Typical markers of objectivity such as freedom from bias, detachment and fact-based reporting date back to the late 19th Century and, despite being deeply ingrained in the Anglo-American news culture, have always been subject to criticism and challenge. Most recently, the growth of openly partisan or populist media has illustrated a deep distrust in traditional news outlets and is overtly questioning whether it is time to jettison objectivity. The increasing use of emotive (and often unfiltered) user-generated content and the rise in citizen journalism appear to have undermined the concept even further. But are we now experiencing a backlash? Through a series of interviews with editorial policy makers at major UK and US news organisations, the paper explores how fake news and other concerns around the impact of social media are leading to fresh debate about objectivity and its potential to make quality journalism stand out

The ϕ6 Cystovirus Protein P7 Becomes Accessible to Antibodies in the Transcribing Nucleocapsid: A Probe for Viral Structural Elements

Protein P7 is a component of the cystovirus viral polymerase complex. In the unpackaged procapsid, the protein is situated in close proximity to the viral directed RNA polymerase, P2. Cryo-electron microscopy difference maps from the species ϕ6 procapsid have demonstrated that P7 and P2 likely interact prior to viral RNA packaging. The location of P7 in the post-packaged nucleocapsid (NC) remains unknown. P7 may translocate closer to the five-fold axis of a filled procapsid but this has not been directly visualized. We propose that monoclonal antibodies (Mabs) can be selected that serve as probe- reagents for viral assembly and structure. A set of Mabs have been isolated that recognize and bind to the ϕ6 P7. The antibody set contains five unique Mabs, four of which recognize a linear epitope and one which recognizes a conformational epitope. The four unique Mabs that recognize a linear epitope display restricted utilization of Vκ and VH genes. The restricted genetic range among 4 of the 5 antibodies implies that the antibody repertoire is limited. The limitation could be the consequence of a paucity of exposed antigenic sites on the ϕ6 P7 surface. It is further demonstrated that within ϕ6 nucleocapsids that are primed for early-phase transcription, P7 is partially accessible to the Mabs, indicating that the nucleocapsid shell (protein P8) has undergone partial disassembly exposing the protein’s antigenic sites

Atrial fibrillation after minimally invasive direct coronary artery bypass surgery

AbstractOBJECTIVESThe study compared the adjusted risk for developing atrial fibrillation (AF) after minimally invasive direct coronary artery bypass surgery (MIDCAB) and coronary artery bypass graft surgery (CABG).BACKGROUNDAtrial fibrillation results in increased morbidity and delays hospital discharge after CABG. Recently, MIDCAB has been explored as an alternative to CABG. Because of differences in surgical approach between the two procedures, the incidence of AF may differ.METHODSRandomly selected patients undergoing CABG and MIDCAB were examined. Baseline variables and postoperative course were recorded through review of medical record data.RESULTSThe MIDCAB patients were younger than CABG patients (64 ± 12 vs. 67 ± 10, p < 0.04) and had less extensive coronary artery disease (53% of MIDCAB vs. 3% of CABG had single-vessel disease, while 15% of MIDCAB vs. 69% of CABG had triple-vessel disease, p < 0.001 for overall group comparisons). No other differences in clinical or treatment data were noted. Postoperative AF occurred less often after MIDCAB (23% vs 39%, p = 0.02). Other significant factors associated with postoperative AF included age (p = 0.0024), prior AF (p = 0.0007), left main disease (p = 0.01), number of vessels bypassed (p = 0.009), absence of postoperative beta-blocker therapy (p = 0.0001), and a serious postoperative complication (p = 0.0018). Because of differences between CABG and MIDCAB patients, multivariate logistic analysis was performed to determine independent predictors of postoperative AF. The type of surgery (CABG vs. MIDCAB) was no longer a significant predictor of postoperative AF (estimated relative risk for AF in CABG vs. MIDCAB patients: 1.57, 95% confidence interval (0.82–2.52).CONCLUSIONSAlthough AF appears to be less common after MIDCAB than after CABG, the lower incidence is due to different clinical characteristics of patients undergoing these procedures

Participatory politics, environmental journalism and newspaper campaigns

This is an Author's Accepted Manuscript of an article published in Journalism Studies, 13(2), 210 - 225, 2012, copyright Taylor & Francis, available online at: http://www.tandfonline.com/10.1080/1461670X.2011.646398.This article explores the extent to which approaches to participatory politics might offer a more useful alternative to understanding the role of environmental journalism in a society where the old certainties have collapsed, only to be replaced by acute uncertainty. This uncertainty not only generates acute public anxiety about risks, it has also undermined confidence in the validity of long-standing premises about the ideal role of the media in society and journalistic professionalism. The consequence, this article argues, is that aspirations of objective reportage are outdated and ill-equipped to deal with many of the new risk stories environmental journalism covers. It is not a redrawing of boundaries that is needed but a wholesale relocation of our frameworks into approaches better suited to the socio-political conditions and uncertainties of late modernity. The exploration of participatory approaches is an attempt to suggest one way this might be done

Coinfection rates in Φ6 bacteriophage are enhanced by virus-induced changes in host cells

Two or more viruses infecting the same host cell can interact in ways that profoundly affect disease dynamics and control, yet the factors determining coinfection rates are incompletely understood. Previous studies have focused on the mechanisms that viruses use to suppress coinfection, but recently the phenomenon of enhanced coinfection has also been documented. In the experiments described here, we explore the hypothesis that enhanced coinfection rates in the bacteriophage Φ6 are achieved by virus-induced upregulation of the Φ6 receptor, which is the bacterial pilus. First, we confirmed that coinfection enhancement in Φ6 is virus-mediated by showing that Φ6 attaches significantly faster to infected cells than to uninfected cells. Second, we explored the hypothesis that coinfection enhancement in Φ6 depends upon changes in the expression of an inducible receptor. Consistent with this hypothesis, the closely related phage, Φ12, that uses constitutively expressed lipopolysaccharide as its receptor, attaches to infected and uninfected cells at the same rate. Our results, along with the previous finding that coinfection in Φ6 is limited to two virions, suggest that viruses may closely regulate rates of coinfection through mechanisms for both coinfection enhancement and exclusion