Cryo-electron tomography of Marburg Virus particles and their morphogenesis within infected cells

Several major human pathogens, including the filoviruses, paramyxoviruses, and rhabdoviruses, package their single-stranded RNA genomes within helical nucleocapsids, which bud through the plasma membrane of the infected cell to release enveloped virions. The virions are often heterogeneous in shape, which makes it difficult to study their structure and assembly mechanisms. We have applied cryo-electron tomography and sub-tomogram averaging methods to derive structures of Marburg virus, a highly pathogenic filovirus, both after release and during assembly within infected cells. The data demonstrate the potential of cryo-electron tomography methods to derive detailed structural information for intermediate steps in biological pathways within intact cells. We describe the location and arrangement of the viral proteins within the virion. We show that the N-terminal domain of the nucleoprotein contains the minimal assembly determinants for a helical nucleocapsid with variable number of proteins per turn. Lobes protruding from alternate interfaces between each nucleoprotein are formed by the C-terminal domain of the nucleoprotein, together with viral proteins VP24 and VP35. Each nucleoprotein packages six RNA bases. The nucleocapsid interacts in an unusual, flexible "Velcro-like" manner with the viral matrix protein VP40. Determination of the structures of assembly intermediates showed that the nucleocapsid has a defined orientation during transport and budding. Together the data show striking architectural homology between the nucleocapsid helix of rhabdoviruses and filoviruses, but unexpected, fundamental differences in the mechanisms by which the nucleocapsids are then assembled together with matrix proteins and initiate membrane envelopment to release infectious virions, suggesting that the viruses have evolved different solutions to these conserved assembly steps.

Pilot evaluation of an adolescent risk and injury prevention programme incorporating curriculum and school connectedness components

School connectedness is an important protective factor for adolescent risk-taking behaviour. This study examined a pilot version of the Skills for Preventing Injury in Youth (SPIY) programme, combining teacher professional development for increasing school connectedness (connectedness component) with a risk and injury prevention curriculum for early adolescents (curriculum component). A process evaluation was conducted on the connectedness component, involving assessments of programme reach, participant receptiveness and initial use, and a preliminary impact evaluation was conducted on the combined connectedness and curriculum programme. The connectedness component was well received by teacher participants, who saw benefits for both themselves and their students. Classroom observation also showed that teachers who received professional development made use of the programme strategies. Grade 8 students who participated in the SPIY programme were less likely to report violent behaviour at six-month follow-up than were control students, and trends also suggested reduced transport injuries. The results of this research support the use of the combined SPIY connectedness and curriculum components in a large-scale effectiveness trial to assess the impact of the programme on students’ connectedness, risk-taking and associated injuries.

Crisis response planning in tunnel and road networks

Critical road infrastructure (such as tunnels and overpasses) is of major significance to society and constitutes major components of interdependent, ‘systems and networks’. Failure in critical components of these wide area infrastructure systems can often result in cascading disturbances with secondary and tertiary impacts - some of which may become initiating sources of failure in their own right, triggering further systems failures across wider networks. Perrow1) considered the impact of our increasing use of technology in high-risk fields, analysing the implications on everyday life and argued that designers of these types of infrastructure systems cannot predict every possible failure scenario nor create perfect contingency plans for operators. Challenges exist for transport system operators in the conceptualisation and implementation of response and subsequent recovery planning for significant events. Disturbances can vary from reduced traffic flow causing traffic congestion throughout the local road network(s) and subsequent possible loss of income to businesses and industry to a major incident causing loss of life or complete loss of an asset. Many organisations and institutions, despite increasing recognition of the effects of crisis events, are not adequately prepared to manage crises2). It is argued that operators of land transport infrastructure are in a similar category of readiness given the recent instances of failures in road tunnels. These unexpected infrastructure failures, and their ultimately identified causes, suggest there is significant room for improvement. As a result, risk profiles for road transport systems are often complex due to the human behaviours and the inter-mix of technical and organisational components and the managerial coverage needed for the socio-technical components and the physical infrastructure. In this sense, the span of managerial oversight may require new approaches to asset management that combines the notion of risk and continuity management. This paper examines challenges in the planning of response and recovery practices of owner/operators of transport systems (above and below ground) in Australia covering: • Ageing or established infrastructure; and • New-build infrastructure. With reference to relevant international contexts this paper seeks to suggest options for enhancing the planning and practice for crisis response in these transport networks and as a result support the resilience of Critical Infrastructure.

Young people, children and their social and spatial citizenship rights

The use of public space by young people and children is a major issue in a number of countries and a range of measures are deployed to to control public space which restrict their social and spatial citizenship rights.

The role of immunoglobulins and their transporters in urogenital Chlamydial infections

Chlamydia trachomatis infections of the male and female reproductive tracts are the world's leading sexually transmitted bacterial disease, and can lead to damaging pathology, scarring and infertility. The resolution of chlamydial infection requires the development of adaptive immune responses to infection, and includes cell-mediated and humoral immunity. Whilst cluster of differentiation (CD)4+ T cells are known to be essential in clearance of infection [1], they are also associated with immune cell infiltration, autoimmunity and infertility in the testes [2-3]. Conversely, antibodies are less associated with inflammation, are readily transported into the reproductive tracts, and can offer lumenal neutralization of chlamydiae prior to infection. Antibodies, or immunoglobulins (Ig), play a supportive role in the resolution of chlamydial infections, and this thesis sought to define the function of IgA and IgG, against a variety of chlamydial antigens expressed during the intracellular and extracellular stages of the chlamydial developmental cycle. Transport of IgA and IgG into the mucosal lumen is facilitated by receptor-mediated transcytosis yet the expression profile (under normal conditions and during urogenital chlamydial infection) of the polymeric immunoglobulin receptor (pIgR) and the neonatal Fc receptor (FcRn) remains unknown. The expression profile of pIgR and FcRn in the murine male reproductive tract was found to be polarized to the lower and upper reproductive tract tissues respectively. This demonstrates that the two receptors have a tissue tropism, which must be considered when targeting pathogens that colonize different sites. In contrast, the expression of pIgR and FcRn in the female mouse was found to be distributed in both the upper and lower reproductive tracts. When urogenitally infected with Chlamydia muridarum, both male and female reproductive tracts up-regulated expression of pIgR and down-regulated expression of FcRn. Unsurprisingly, the up-regulation of pIgR increased the concentration of IgA in the lumen. However, down-regulation of FcRn, prevented IgG uptake and led to an increase or pooling of IgG in lumenal secretions. As previous studies have identified the importance of pIgR-mediated delivery of IgA, as well as the potential of IgA to bind and neutralize intracellular pathogens, IgA against a variety of chlamydial antigens was investigated. The protection afforded by IgA against the extracellular antigen major outer membrane protein (MOMP), was found to be dependent on pIgR expression in vitro and in vivo. It was also found that in the absence of pIgR, no protection was afforded to mice previously immunized with MOMP. The protection afforded from polyclonal IgA against the intracellular chlamydial antigens; inclusion membrane protein A (IncA), inclusion membrane proteins (IncMem) and secreted chlamydial protease-like activity factor (CPAF) were produced and investigated in vitro. Antigen-specific intracellular IgA was found to bind to the respective antigen within the infected cell, but did not significantly reduce inclusion formation (p > 0.05). This suggests that whilst IgA specific for the selected antigens was transported by pIgR to the chlamydial inclusion, it was unable to prevent growth. Similarly, immunization of male mice with intracellular chlamydial antigens (IncA or IncMem), followed by depletion CD4+ T cells, and subsequent urogenital C. muridarum challenge, provided minimal pIgR-mediated protection. Wild type male mice immunized with IncA showed a 57 % reduction (p < 0.05), and mice deficient in pIgR showed a 35 % reduction (p < 0.05) in reproductive tract chlamydial burden compared to control antigen, and in the absence of CD4+ T cells. This suggests that pIgR and secretory IgA (SIgA) were playing a protective role (21 % pIgR-mediated) in unison with another antigen-specific immune mechanism (36 %). Interestingly, IgA generated during a primary respiratory C. muridarum infection did not provide a significant amount of protection to secondary urogenital C. muridarum challenge. Together, these data suggest that IgA specific for an extracellular antigen (MOMP) can play a strong protective role in chlamydial infections, and that IgA targeting intracellular antigens is also effective but dependent on pIgR expression in tissues. However, whilst not investigated here, IgA targeting and blocking other intracellular chlamydial antigens, that are more essential for replication or type III secretion, may be more efficacious in subunit vaccines. Recently, studies have demonstrated that IgG can neutralize influenza virus by trafficking IgG-bound virus to lysosomes [4]. We sought to determine if this process could also traffic chlamydial antigens for degradation by lysosomes, despite Chlamydia spp. actively inhibiting fusion with the host endocytic pathway. As observed in pIgR-mediated delivery of anti-IncA IgA, FcRn similarly transported IgG specific for IncA which bound the inclusion membrane. Interestingly, FcRn-mediated delivery of anti-IncA IgG significantly decreased inclusion formation by 36 % (p < 0.01), and induced aberrant inclusion morphology. This suggests that unlike IgA, IgG can facilitate additional host cellular responses which affect the intracellular niche of chlamydial growth. Fluorescence microscopy revealed that IgG also bound the inclusion, but unlike influenza studies, did not induce the recruitment of lysosomes. Notably, anti-IncA IgG recruited sequestosomes to the inclusion membrane, markers of the ubiquitin/proteasome pathway and major histocompatibility complex (MHC) class I loading. To determine if the protection against C. muridarum infection afforded by IncA IgG in vitro translated in vivo, wild type mice and mice deficient in functional FcRn and MHC-I, were immunized, depleted of CD4+, and urogenitally infected with C. muridarum. Unlike in pIgR-deficient mice, the protection afforded from IncA immunization was completely abrogated in mice lacking functional FcRn and MHC-I/CD8+. Thus, both anti-IncA IgA and IgG can bind the inclusion in a pIgR and FcRn-mediated manner, respectively. However, only IgG mediates a higher reduction in chlamydial infection in vitro and in vivo suggesting more than steric blocking of IncA had occurred. Unlike anti-MOMP IgA, which reduced chlamydial infection of epithelial cells and male mouse tissues, IgG was found to enhance infectivity in vitro, and in vivo. Opsonization of EBs with MOMP-IgG enhanced inclusion formation of epithelial cells in a MOMP-IgG dose-dependent and FcRn-dependent manner. When MOMP-IgG opsonized EBs were inoculated into the vagina of female mice, a small but non-significant (p > 0.05) enhancement of cervicovaginal C. muridarum shedding was observed three days post infection in mice with functional FcRn. Interestingly, infection with opsonized EBs reduced the intensity of the peak of infection (day six) but protracted the duration of infection by 60 % in wild type mice only. Infection with EBs opsonized in IgG also significantly increased (p < 0.05) hydrosalpinx formation in the oviducts and induced lymphocyte infiltration uterine horns. As MOMP is an immunodominant antigen, and is widely used in vaccines, the ability of IgG specific to extracellular chlamydial antigens to enhance infection and induce pathology needs to be considered. Together, these data suggest that immunoglobulins play a dichotomous role in chlamydial infections, and are dependent on antigen specificity, FcRn and pIgR expression. FcRn was found to be highly expressed in upper male reproductive tract, whilst pIgR was dominantly expressed in the lower reproductive tract. Conversely, female mice expressed FcRn and pIgR in both the lower and upper reproductive tracts. In response to a normal chlamydial infection, pIgR is up-regulated increasing secretory IgA release, but FcRn is down-regulated preventing IgG uptake. Similarly to other studies [5-6], we demonstrate that IgA and IgG generated during primary chlamydial infections plays a minor role in recall immunity, and that antigen-specific subunit vaccines can offer more protection. We also show that both IgA and IgG can be used to target intracellular chlamydial antigens, but that IgG is more effective. Finally, IgA against the extracellular antigen MOMP can afford protection, whist IgG plays a deleterious role by increasing infectivity and inducing damaging immunopathology. Further investigations with additional antigens or combination subunit vaccines will enhance our understanding the protection afforded by antibodies against intracellular and extracellular pathogenic antigens, and help improve the development of an efficacious chlamydial vaccine.

Simulating land cover changes and their impacts on land surface temperature in Dhaka, Bangladesh

Despite research that has been conducted elsewhere, little is known, to-date, about land cover dynamics and their impacts on land surface temperature (LST) in fast growing mega cities of developing countries. Landsat satellite images of 1989, 1999, and 2009 of Dhaka Metropolitan (DMP) area were used for analysis. This study first identified patterns of land cover changes between the periods and investigated their impacts on LST; second, applied artificial neural network to simulate land cover changes for 2019 and 2029; and finally, estimated their impacts on LST in respective periods. Simulation results show that if the current trend continues, 56% and 87% of the DMP area will likely to experience temperatures in the range of greater than or equal to 30°C in 2019 and 2029, respectively. The findings possess a major challenge for urban planners working in similar contexts. However, the technique presented in this paper would help them to quantify the impacts of different scenarios (e.g., vegetation loss to accommodate urban growth) on LST and consequently to devise appropriate policy measures.

Modeling users’ web search behavior and their cognitive styles

Previous studies have shown that users’ cognitive styles play an important role during Web searching. However, only limited studies have showed the relationship between cognitive styles and Web search behavior. Most importantly, it is not clear which components of Web search behavior are influenced by cognitive styles. This paper examines the relationships between users’ cognitive styles and their Web searching and develops a model that portrays the relationship. The study uses qualitative and quantitative analyses to inform the study results based on data gathered from 50 participants. A questionnaire was utilised to collect participants’ demographic information, and Riding’s (1991) Cognitive Style Analysis (CSA) test to assess their cognitive styles. Results show that users’ cognitive styles influenced their information searching strategies, query reformulation behaviour, Web navigational styles and information processing approaches. The user model developed in this study depicts the fundamental relationships between users’ Web search behavior and their cognitive styles. Modeling Web search behavior with a greater understanding of user’s cognitive styles can help information science researchers and information systems designers to bridge the semantic gap between the user and the systems. Implications of the research for theory and practice, and future work are discussed.

Expression patterns of vascular-specific promoters RolC and Sh in transgenic potatoes and their use in engineering PLRV-resistant plants

The expression patterns of GUS fusion constructs driven by the Agrobacterium rhizogenes RolC and the maize Sh (Shrunken: sucrose synthase-1) promoters were examined in transgenic potatoes (cv. Atlantic). RolC drove high-level gene expression in phloem tissue, bundle sheath cells and vascular parenchyma, but not in xylem or non-vascular tissues. Sh expression was exclusively confined to phloem tissue. Potato leafroll luteovirus (PLRV) replicates only in phloem tissues, and we show that when RolC is used to drive expression of the PLRV coat protein gene, virus-resistant lines can be obtained. In contrast, no significant resistance was observed when the Sh promoter was used.

Eph receptors and their ligands : promising molecular biomarkers and therapeutic targets in prostate cancer

Although at present, there is a high incidence of prostate cancer, particularly in the Western world, mortality from this disease is declining and occurs primarily only from clinically significant late stage tumors with a poor prognosis. A major current focus of this field is the identification of new biomarkers which can detect earlier, and more effectively, clinically significant tumors from those deemed “low risk”, as well as predict the prognostic course of a particular cancer. This strategy can in turn offer novel avenues for targeted therapies. The large family of Receptor Tyrosine Kinases, the Ephs, and their binding partners, the ephrins, has been implicated in many cancers of epithelial origin through stimulation of oncogenic transformation, tumor angiogenesis, and promotion of increased cell survival, invasion and migration. They also show promise as both biomarkers of diagnostic and prognostic value and as targeted therapies in cancer. This review will briefly discuss the complex roles and biological mechanisms of action of these receptors and ligands and, with regard to prostate cancer, highlight their potential as biomarkers for both diagnosis and prognosis, their application as imaging agents, and current approaches to assessing them as therapeutic targets. This review demonstrates the need for future studies into those particular family members that will prove helpful in understanding the biology and potential as targets for treatment of prostate cancer.

Study Protocol - Alcohol Management Plans (AMPs) in remote indigenous communities in Queensland: their impacts on injury, violence, health and social indicators and their cost-effectiveness

Background In 2002/03 the Queensland Government responded to high rates of alcohol-related harm in discrete Indigenous communities by implementing alcohol management plans (AMPs), designed to include supply and harm reduction and treatment measures. Tighter alcohol supply and carriage restrictions followed in 2008 following indications of reductions in violence and injury. Despite the plans being in place for over a decade, no comprehensive independent review has assessed to what level the designed aims were achieved and what effect the plans have had on Indigenous community residents and service providers. This study will describe the long-term impacts on important health, economic and social outcomes of Queensland’s AMPs. Methods/Design The project has two main studies, 1) outcome evaluation using de-identified epidemiological data on injury, violence and other health and social indicators for across Queensland, including de-identified databases compiled from relevant routinely-available administrative data sets, and 2) a process evaluation to map the nature, timing and content of intervention components targeting alcohol. Process evaluation will also be used to assess the fidelity with which the designed intervention components have been implemented, their uptake and community responses to them and their perceived impacts on alcohol supply and consumption, injury, violence and community health. Interviews and focus groups with Indigenous residents and service providers will be used. The study will be conducted in all 24 of Queensland’s Indigenous communities affected by alcohol management plans. Discussion This evaluation will report on the impacts of the original aims for AMPs, what impact they have had on Indigenous residents and service providers. A central outcome will be the establishment of relevant databases describing the parameters of the changes seen. This will permit comprehensive and rigorous surveillance systems to be put in place and provided to communities empowering them with the best credible evidence to judge future policy and program requirements for themselves. The project will inform impending alcohol policy and program adjustments in Queensland and other Australian jurisdictions. The project has been approved by the James Cook University Human Research Ethics Committee (approval number H4967 & H5241).

Pharmaceutical company influences on medication prescribing and their potential impact on quality use of medicines

Background Pharmaceuticals are big business, reporting strong market growth year after year. The ‘gatekeepers’ of this market are prescribers of medicines, who are the major target of pharmaceutical companies, utilizing direct and indirect influences. Methods This paper draws on previous research investigating pharmaceutical company prescribing influences to develop a qualitative model demonstrating the synergism between commercial influences on prescribing. The generic model was used to explore a realistic but hypothetical scenario to ascertain the applicability of the model. Results and Discussion A generic influence model was developed. The model was readily able to be adapted to reflect a realistic practice scenario. Conclusion Prescriber awareness of the linkages between various seemingly separate marketing techniques could potentially improve medicines usage in an evidence-based practice paradigm.

Functional MRI of facial emotion recognition deficits in schizophrenia and their electrophysiological correlates

Empirical evidence suggests impaired facial emotion recognition in schizophrenia. However, the nature of this deficit is the subject of ongoing research. The current study tested the hypothesis that a generalized deficit at an early stage of face-specific processing (i.e. putatively subserved by the fusiform gyrus) accounts for impaired facial emotion recognition in schizophrenia as opposed to the Negative Emotion-specific Deficit Model, which suggests impaired facial information processing at subsequent stages. Event-related potentials (ERPs) were recorded from 11 schizophrenia patients and 15 matched controls while performing a gender discrimination and a facial emotion recognition task. Significant reduction of the face-specific vertex positive potential (VPP) at a peak latency of 165 ms was confirmed in schizophrenia subjects whereas their early visual processing, as indexed by P1, was found to be intact. Attenuated VPP was found to correlate with subsequent P3 amplitude reduction and to predict accuracy when performing a facial emotion discrimination task. A subset of ten schizophrenia patients and ten matched healthy control subjects also performed similar tasks in the magnetic resonance imaging scanner. Patients showed reduced blood oxygenation level-dependent (BOLD) activation in the fusiform, inferior frontal, middle temporal and middle occipital gyrus as well as in the amygdala. Correlation analyses revealed that VPP and the subsequent P3a ERP components predict fusiform gyrus BOLD activation. These results suggest that problems in facial affect recognition in schizophrenia may represent flow-on effects of a generalized deficit in early visual processing.

Novel loci affecting iron homeostasis and their effects in individuals at risk for hemochromatosis

Variation in body iron is associated with or causes diseases, including anaemia and iron overload. Here, we analyse genetic association data on biochemical markers of iron status from 11 European-population studies, with replication in eight additional cohorts (total up to 48,972 subjects). We find 11 genome-wide-significant (P<5 × 10−8) loci, some including known iron-related genes (​HFE, ​SLC40A1, ​TF, ​TFR2, ​TFRC, ​TMPRSS6) and others novel (​ABO, ​ARNTL, ​FADS2, ​NAT2, ​TEX14). SNPs at ​ARNTL, ​TF, and ​TFR2 affect iron markers in ​HFE C282Y homozygotes at risk for hemochromatosis. There is substantial overlap between our iron loci and loci affecting erythrocyte and lipid phenotypes. These results will facilitate investigation of the roles of iron in disease.

The trajectory of minor stroke recovery for men and their female spousal caregivers: Literature review

Title The trajectory of minor stroke recovery for men and their female spousal caregivers: literature review Aim This paper is a report of a narrative review to examine the current state of knowledge regarding the impact of minor stroke on male patients and their female spousal caregivers’ recovery trajectory and quality of life. Background Minor stroke survivors are often discharged early in the recovery process. The perception of the healthcare community that these patients and their female spousal caregivers will experience an uneventful recovery may lead to inadequate preparation for the postdischarge period. Methods A range of databases was searched to identify papers addressing ‘minor stroke’, ‘transitions’, ‘quality of life’, ‘chronic disease’, ‘caregivers’ and ‘spouse caregivers’, including AARP Ageline, AMED, CINAHL, Evidence Based Medicine Reviews, MEDLINE and PsychInfo. Papers published in English from 1990 to December 2006 were included. Thirty-four papers were in the final data set. Results Minor stroke survivors and their female spousal caregivers may experience major challenges in adaptations postdischarge. The trajectory of minor stroke recovery may necessitate a re-evaluation of life plans, rethinking of priorities and integration of resulting disabilities into current and emerging life situations for both stroke survivors and their female spousal caregivers. In many cases these adaptations are compounded by transitions associated with the normal ageing process. Conclusion While there is extensive literature on stroke recovery and the role of caregivers in general, there is little available describing the recovery of minor stroke survivors in relation to the normal ageing process. Further research is needed examining recovery from a transitional perspective, to support nurses and other health professionals discharge planning.