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Munday D.C.,University of Liverpool | Wu W.,University of Liverpool | Smith N.,Roslin Institute | Fix J.,French National Institute for Agricultural Research | And 18 more authors.
Journal of Virology | Year: 2015

The human respiratory syncytial virus (HRSV) core viral RNA polymerase comprises the large polymerase protein (L) and its cofactor, the phosphoprotein (P), which associate with the viral ribonucleoprotein complex to replicate the genome and, together with the M2-1 protein, transcribe viral mRNAs. While cellular proteins have long been proposed to be involved in the synthesis of HRSV RNA by associating with the polymerase complex, their characterization has been hindered by the difficulty of purifying the viral polymerase from mammalian cell culture. In this study, enhanced green fluorescent protein (EGFP)-tagged L- and P-protein expression was coupled with high-affinity anti-GFP antibody-based immunoprecipitation and quantitative proteomics to identify cellular proteins that interacted with either the L- or the P-proteins when expressed as part of a biologically active viral RNP. Several core groups of cellular proteins were identified that interacted with each viral protein including, in both cases, protein chaperones. Ablation of chaperone activity by using small-molecule inhibitors confirmed previously reported studies which suggested that this class of proteins acted as positive viral factors. Inhibition of HSP90 chaperone function in the current study showed that HSP90 is critical for L-protein function and stability, whether in the presence or absence of the P-protein. Inhibition studies suggested that HSP70 also disrupts virus biology and might help the polymerase remodel the nucleocapsid to allow RNA synthesis to occur efficiently. This indicated a proviral role for protein chaperones in HRSV replication and demonstrates that the function of cellular proteins can be targeted as potential therapeutics to disrupt virus replication. © 2015, American Society for Microbiology.


Read J.M.,University of Liverpool | Read J.M.,NIHR Health Protection Research Unit in Emerging and Zoonotic Infections | Smith V.,London School of Hygiene and Tropical Medicine | Gething P.W.,University of Oxford | And 2 more authors.
Malaria Journal | Year: 2014

Background: An increasing proportion of malaria cases diagnosed in UK residents with a history of travel to malaria endemic areas are due to Plasmodium falciparum. Methods. In order to identify travellers at most risk of acquiring malaria a proportional hazards model was used to estimate the risk of acquiring malaria stratified by purpose of travel and age whilst adjusting for entomological inoculation rate (EIR) and duration of stay in endemic countries. Results: Travellers visiting friends and relatives and business travellers were found to have significantly higher hazard of acquiring malaria (adjusted hazard ratio (HR) relative to that of holiday makers 7.4, 95% CI 6.4-8.5, p < 0. 0001 and HR 3.4, 95% CI 2.9-3.8, p < 0. 0001, respectively). All age-groups were at lower risk than children aged 0-15 years. Conclusions: These estimates of the increased risk for business travellers and those visiting friends and relatives should be used to inform programmes to improve awareness of the risks of malaria when travelling. © 2014 Pinsent et al.; licensee BioMed Central Ltd.


Griffiths M.J.,University of Liverpool | Griffiths M.J.,Alder Hey Childrens NHS Foundation Trust | Turtle L.,University of Liverpool | Solomon T.,University of Liverpool | And 2 more authors.
Handbook of Clinical Neurology | Year: 2014

Japanese encephalitis is the most important epidemic encephalitis in the world, responsible for a higher burden of disability than any other arthropod-borne virus. Twenty-four countries, home to over 2.5 billion people, within Asia and the Western Pacific rim are endemic for Japanese encephalitis. In these countries the burden of disease is mostly on children. The adult population is typically immune following childhood exposure. Although only a minority of people exposed to the virus develop disease, Japanese encephalitis is estimated to be responsible for nearly 70 000 cases and 20 000 deaths annually. Patients develop a febrile illness followed by headache, vomiting, impaired consciousness, and often seizures. A wide range of neurologic abnormalities are observed. Typically 20-30% of patients die, and 30-50% of survivors have severe neuropsychiatric sequelae. There are no proven effective therapies and treatment is supportive; however, Japanese encephalitis can be prevented by vaccination. © 2014 Elsevier B.V.


Broadhurst M.J.,Partners in Health | Kelly J.D.,Partners in Health | Miller A.,Partners in Health | Miller A.,Harvard University | And 23 more authors.
The Lancet | Year: 2015

Background At present, diagnosis of Ebola virus disease requires transport of venepuncture blood to field biocontainment laboratories for testing by real-time RT-PCR, resulting in delays that complicate patient care and infection control efforts. Therefore, an urgent need exists for a point-of-care rapid diagnostic test for this disease. In this Article, we report the results of a field validation of the Corgenix ReEBOV Antigen Rapid Test kit. Methods We performed the rapid diagnostic test on fingerstick blood samples from 106 individuals with suspected Ebola virus disease presenting at two clinical centres in Sierra Leone. Adults and children who were able to provide verbal consent or assent were included; we excluded patients with haemodynamic instability and those who were unable to cooperate with fingerstick or venous blood draw. Two independent readers scored each rapid diagnostic test, with any disagreements resolved by a third. We compared point-of-care rapid diagnostic test results with clinical real-time RT-PCR results (RealStar Filovirus Screen RT-PCR kit 1·0; altona Diagnostics GmbH, Hamburg, Germany) for venepuncture plasma samples tested in a Public Health England field reference laboratory (Port Loko, Sierra Leone). Separately, we performed the rapid diagnostic test (on whole blood) and real-time RT-PCR (on plasma) on 284 specimens in the reference laboratory, which were submitted to the laboratory for testing from many clinical sites in Sierra Leone, including our two clinical centres. Findings In point-of-care testing, all 28 patients who tested positive for Ebola virus disease by RT-PCR were also positive by fingerstick rapid diagnostic test (sensitivity 100% [95% CI 87·7-100]), and 71 of 77 patients who tested negative by RT-PCR were also negative by the rapid diagnostic test (specificity 92·2% [95% CI 83·8-97·1]). In laboratory testing, all 45 specimens that tested positive by RT-PCR were also positive by the rapid diagnostic test (sensitivity 100% [95% CI 92·1-100]), and 214 of 232 specimens that tested negative by RT-PCR were also negative by the rapid diagnostic test (specificity 92·2% [88·0-95·3]). The two independent readers agreed about 95·2% of point-of-care and 98·6% of reference laboratory rapid diagnostic test results. Cycle threshold values ranged from 15·9 to 26·3 (mean 22·6 [SD 2·6]) for the PCR-positive point-of-care cohort and from 17·5 to 26·3 (mean 21·5 [2·7]) for the reference laboratory cohort. Six of 16 banked plasma samples from rapid diagnostic test-positive and altona-negative patients were positive by an alternative real-time RT-PCR assay (the Trombley assay); three (17%) of 18 samples from individuals who were negative by both the rapid diagnostic test and altona test were also positive by Trombley. Interpretation The ReEBOV rapid diagnostic test had 100% sensitivity and 92% specificity in both point-of-care and reference laboratory testing in this population (maximum cycle threshold 26·3). With two independent readers, the test detected all patients who were positive for Ebola virus by altona real-time RT-PCR; however, this benchmark itself had imperfect sensitivity. Funding Abundance Foundation. © 2015 Elsevier Ltd.


Fisher D.L.,Queen Mary, University of London | Defres S.,University of Liverpool | Solomon T.,University of Liverpool | Solomon T.,NIHR Health Protection Research Unit in Emerging and Zoonotic Infections | Solomon T.,The Walton Center Neurology Foundation Trust
QJM | Year: 2015

Encephalitis is the most frequent neurological complication of measles virus infection. This review examines the pathophysiology of measles infection and the presentations, diagnosis and treatment of the four types of measles-induced encephalitis including primary measles encephalitis, acute post-measles encephalitis, measles inclusion body encephalitis and subacute sclerosing panencephalitis. The early symptoms of encephalitis may be non-specific and can be mistakenly attributed to a systemic infection leading to a delay in diagnosis. This review provides a summary of the symptoms that should cause health care workers to suspect measles-induced encephalitis. © The Author 2014.


Garcia-Dorival I.,University of Liverpool | Garcia-Dorival I.,NIHR Health Protection Research Unit in Emerging and Zoonotic Infections | Wu W.,University of Liverpool | Dowall S.,Public Health England | And 13 more authors.
Journal of Proteome Research | Year: 2014

Viral pathogenesis in the infected cell is a balance between antiviral responses and subversion of host-cell processes. Many viral proteins specifically interact with host-cell proteins to promote virus biology. Understanding these interactions can lead to knowledge gains about infection and provide potential targets for antiviral therapy. One such virus is Ebola, which has profound consequences for human health and causes viral hemorrhagic fever where case fatality rates can approach 90%. The Ebola virus VP24 protein plays a critical role in the evasion of the host immune response and is likely to interact with multiple cellular proteins. To map these interactions and better understand the potential functions of VP24, label-free quantitative proteomics was used to identify cellular proteins that had a high probability of forming the VP24 cellular interactome. Several known interactions were confirmed, thus placing confidence in the technique, but new interactions were also discovered including one with ATP1A1, which is involved in osmoregulation and cell signaling. Disrupting the activity of ATP1A1 in Ebola-virus-infected cells with a small molecule inhibitor resulted in a decrease in progeny virus, thus illustrating how quantitative proteomics can be used to identify potential therapeutic targets. © 2014 American Chemical Society.


Liu L.,Northwest University, China | Lear Z.,University of Leeds | Hughes D.J.,University of Leeds | Wu W.,University of Liverpool | And 6 more authors.
Veterinary Microbiology | Year: 2015

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major threat to the swine industry and food security worldwide. The nucleocapsid (N) protein is a major structural protein of PRRSV. The primary function of this protein is to encapsidate the viral RNA genome, and it is also thought to participate in the modulation of host cell biology and recruitment of cellular factors to facilitate virus infection. In order to the better understand these latter roles the cellular interactome of PRRSV N protein was defined using label free quantitative proteomics. This identified several cellular factors that could interact with the N protein including poly [ADP-ribose] polymerase 1 (PARP-1), a cellular protein, which can add adenosine diphosphate ribose to a protein. Use of the PARP-1 small molecule inhibitor, 3-AB, in PRRSV infected cells demonstrated that PARP-1 was required and acted as an enhancer factor for virus biology. Serial growth of PRRSV in different concentrations of 3-AB did not yield viruses that were able to grow with wild type kinetics, suggesting that by targeting a cellular protein crucial for virus biology, resistant phenotypes did not emerge. This study provides further evidence that cellular proteins, which are critical for virus biology, can also be targeted to ablate virus growth and provide a high barrier for the emergence of drug resistance. © 2014 Elsevier B.V.


Caminade C.,University of Liverpool | van Dijk J.,University of Liverpool | Baylis M.,University of Liverpool | Baylis M.,NIHR Health Protection Research Unit in Emerging and Zoonotic Infections | Williams D.,University of Liverpool
Geospatial Health | Year: 2015

Fasciola hepatica is a parasitic worm responsible for fasciolosis in grazed ruminants in Europe. The free-living stages of this parasite are sensitive to temperature and soil moisture, as are the intermediate snail hosts the parasite depends on for its life-cycle. We used a climate-driven disease model in order to assess the impact of recent and potential future climate changes on the incidence of fasciolosis and to estimate the related uncertainties at the scale of the European landmass. The current climate appears to be highly suitable for fasciolosis throughout the European Union with the exception of some parts of the Mediterranean region. Simulated climatic suitability for fasciolosis significantly increased during the 2000s in central and north-western Europe, which is consistent with an observed increased in ruminant infections. The simulation showed that recent trends are likely to continue in the future with the estimated pattern of climate change for northern Europe, possibly extending the season suitable for development of the parasite in the environment by up to four months. For southern Europe, the simulated burden of disease may be lower, but the projected climate change will increase the risk during the winter months, since the simulated changes in temperature and moisture support the development of the free-living and intra-molluscan stages between November and March. In the event of predicted climate change, F. hepatica will present a serious risk to the health, welfare and productivity of all ruminant livestock. Improved, bespoke control programmes, both at farm and region levels, will then become imperative if problems, such as resistance of the parasite associated with increased drug use, are to be mitigated. © 2015, Page Press Publications. All rights reserved.


Awad F.,Institute of Infection and Global Health | Awad F.,Omar Al-Mukhtar University | Hutton S.,Institute of Infection and Global Health | Forrester A.,Institute of Infection and Global Health | And 3 more authors.
Avian Pathology | Year: 2016

Groups of one-day-old broiler chicks were vaccinated via the oculo-nasal route with different live infectious bronchitis virus (IBV) vaccines: Massachusetts (Mass), 793B, D274 or Arkansas (Ark). Clinical signs and gross lesions were evaluated. Five chicks from each group were humanely killed at intervals and their tracheas collected for ciliary activity assessment and for the detection of CD4+, CD8+ and IgA-bearing B cells by immunohistochemistry (IHC). Blood samples were collected at intervals for the detection of anti-IBV antibodies. At 21 days post-vaccination (dpv), protection conferred by different vaccination regimes against virulent M41, QX and 793B was assessed. All vaccination programmes were able to induce high levels of CD4+, CD8+ and IgA-bearing B cells in the trachea. Significantly higher levels of CD4+ and CD8+ expression were observed in the Mass2+ 793B2-vaccinated group compared to the other groups (subscripts indicate different manufacturers). Protection studies showed that the group of chicks vaccinated with Mass2+ 793B2 produced 92% ciliary protection against QX challenge; compared to 53%, 68% and 73% ciliary protection against the same challenge virus by Mass1+ D274, Mass1+ 793B1 and Mass3+ Ark, respectively. All vaccination programmes produced more than 85% ciliary protection against M41 and 793B challenges. It appears that the variable levels of protection provided by different heterologous live IBV vaccinations are dependent on the levels of local tracheal immunity induced by the respective vaccine combination. The Mass2+ 793B2 group showed the worst clinical signs, higher mortality and severe lesions following vaccination, but had the highest tracheal immune responses and demonstrated the best protection against all three challenge viruses. © 2016 Houghton Trust Ltd.


Westgarth C.,University of Liverpool | Christley R.M.,University of Liverpool | Christley R.M.,NIHR Health Protection Research Unit in Emerging and Zoonotic Infections | Christian H.E.,University of Western Australia
International Journal of Behavioral Nutrition and Physical Activity | Year: 2014

Background: Physical inactivity and sedentary behaviour are major threats to population health. A considerable proportion of people own dogs, and there is good evidence that dog ownership is associated with higher levels of physical activity. However not all owners walk their dogs regularly. This paper comprehensively reviews the evidence for correlates of dog walking so that effective interventions may be designed to increase the physical activity of dog owners.Methods: Published findings from 1990-2012 in both the human and veterinary literature were collated and reviewed for evidence of factors associated with objective and self-reported measures of dog walking behaviour, or reported perceptions about dog walking. Study designs included cross-sectional observational, trials and qualitative interviews.Results: There is good evidence that the strength of the dog-owner relationship, through a sense of obligation to walk the dog, and the perceived support and motivation a dog provides for walking, is strongly associated with increased walking. The perceived exercise requirements of the dog may also be a modifiable point for intervention. In addition, access to suitable walking areas with dog supportive features that fulfil dog needs such as off-leash exercise, and that also encourage human social interaction, may be incentivising.Conclusion: Current evidence suggests that dog walking may be most effectively encouraged through targeting the dog-owner relationship and by providing dog-supportive physical environments. More research is required to investigate the influence of individual owner and dog factors on 'intention' to walk the dog as well as the influence of human social interaction whilst walking a dog. The effects of policy and cultural practices relating to dog ownership and walking should also be investigated. Future studies must be of a higher quality methodological design, including accounting for the effects of confounding between variables, and longitudinal designs and testing of interventions in a controlled design in order to infer causality. © 2014 Westgarth et al.; licensee BioMed Central Ltd.

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