A Two-Phase Innate Host Response to Alphavirus Infection Identified by mRNP-Tagging In Vivo

A concept fundamental to viral pathogenesis is that infection induces specific changes within the host cell, within specific tissues, or within the entire animal. These changes are reflected in a cascade of altered transcription patterns evident during infection. However, elucidation of this cascade in vivo has been limited by a general inability to distinguish changes occurring in the minority of infected cells from those in surrounding uninfected cells. To circumvent this inherent limitation of traditional gene expression profiling methods, an innovative mRNP-tagging technique was implemented to isolate host mRNA specifically from infected cells in vitro as well as in vivo following Venezuelan equine encephalitis virus (VEE) infection. This technique facilitated a direct characterization of the host defense response specifically within the first cells infected with VEE, while simultaneous total RNA analysis assessed the collective response of both the infected and uninfected cells. The result was a unique, multifaceted profile of the early response to VEE infection in primary dendritic cells, as well as in the draining lymph node, the initially targeted tissue in the mouse model. A dynamic environment of complex interactions was revealed, and suggested a two-step innate response in which activation of a subset of host genes in infected cells subsequently leads to activation of the surrounding uninfected cells. Our findings suggest that the application of viral mRNP-tagging systems, as introduced here, will facilitate a much more detailed understanding of the highly coordinated host response to infectious agents

Alphavirus replicon-based enhancement of mucosal and systemic immunity is linked to the innate response generated by primary immunization

Venezuelan equine encephalitis virus replicon particles (VRP) function as an effective systemic, cellular and mucosal adjuvant when codelivered with antigen, and show promise for use as a component in new and existing human vaccine formulations. We show here that VRP are effective at low dose and by intramuscular delivery, two useful features for implementation of VRP as a vaccine adjuvant. In mice receiving a prime and boost with antigen, we found that VRP are required in prime only to produce a full adjuvant effect. This outcome indicates that the events triggered during prime with VRP are sufficient to establish the nature and magnitude of the immune response to a second exposure to antigen. Events induced by VRP in the draining lymph node after prime include robust secretion of many inflammatory cytokines, upregulation of CD69 on leukocytes, and increased cellularity, with a disproportionate increase of a cell population expressing CD11c, CD11b, and F4/80. We show that antigen delivered 24 hours after administration of VRP does not benefit from an adjuvant effect, indicating that the events which are critical to VRP-mediated adjuvant activity occur within the first 24 hours. Further studies of the events induced by VRP will help elucidate the mechanism of VRP adjuvant activity and will advance the safe implementation of this adjuvant in human vaccines

Crop Updates 2002 - Farming Systems

This session covers forty one papers from different authors: INTRODUCTION 1. Future Farming Systems session for Crop Updates 2002 Peter Metcalf, FARMING SYSTEMS SUBPROGRAM MANAGER GRAINS PROGRAM Department of Agriculture 2. Perennial pastures in annual cropping systems: Lucerne and beyond, the ‘Big Picture’, Mike Ewing, Deputy CEO CRC for Plant-based Management of Dryland Salinity, Department of Agriculture 3. Perennial pastures in annual cropping systems: lucerne and beyond, Roy Latta and Keith Devenish, Department of Agriculture 4. Establishing Lucerne with a cover crop, Diana Fedorenko1, Clayton Butterly1, Chantelle Butterly1, Kim and Neil Diamond2, Stuart McAlpine2, Bill Bowden1, Jessica Johns3, 1Centre for Cropping Systems, Northam, 2Farmer, Buntine, 3Department of Agriculture 5. Overcropping: Chemical suppression of Lucerne, Terry Piper1, Diana Fedorenko1, Clayton Butterly1, Chantelle Butterly1, Stuart McAlpine2, Jessica Johns3, 1Centre for Cropping Systems, Northam, 2Farmer, Buntine, 3Department of Agriculture 6. Overcropping: Effect of Lucerne density on crop yield, Diana Fedorenko1, Bill Bowden1, Clayton Butterly1, Chantelle Butterly1, Stuart McAlpine2, Terry Piper1,1Centre for Cropping Systems, Department of Agriculture, Northam, 2Farmer, Buntine 7. Residual effect of weed management in the third year of Lucerne on the following wheat crop, Diana Fedorenko1, Clayton Butterly1, Chantelle Butterly1, Stuart McAlpine2,Terry Piper1, David Bowran1, Jessica Johns3,1Centre for Cropping Systems, Northam, 2Farmer, Buntine, 3Department of Agriculture 8. Production of Lucerne and serradella in four soil types, Diana Fedorenko1 Clayton Butterly1, Chantelle Butterly1, Robert Beard2 1Centre for Cropping Systems, Department of Agriculture, 2Farmer, Cunderdin 9. The effect of spray topping on newly established Lucerne, Keith Devenish, Agriculture Western Australia 10. Leakage from phase rotations involving Lucerne, Phil Ward, CSIRO Plant Industry 11. Fungal diseases present in Western Australian Lucerne crops, Dominie Wright and Nichole Burges, Department of Agriculture 12. Survey of Western Australian Lucerne stands reveals widespread virus infection, Roger Jones and Danae Harman, Crop Improvement Institute, Department of Agriculture, and Centre for Legumes in Mediterranean Agriculture, University of WA ANNUAL PASTURE SYSTEMS 13. The use of Twist Fungus as a biosecurity measure against Annual Ryegrass Toxicity (ARGT), Greg Shea, GrainGuard Coordinator and George Yan, Biological and Resource Technology 14.Limitations and opportunities for increasing water use by annual crops and pastures, David Tennant1, Phil Ward2and David Hall1 1Department of Agriculture, 2CSIRO, Plant Industries, Floreat Park 15. Developing pasture species mixtures for more productive and sustainable cropping systems – 2001 crop performance, Anyou Liu, Clinton Revell and Candy Hudson, Centre for Cropping Systems, Department of Agriculture 16. Developing pasture species mixtures for more productive and sustainable cropping systems – weed management in regenerating mixtures, Anyou Liu and Clinton Revell, Centre for Cropping Systems, Department of Agriculture 17. Aphid tolerance of annual pasture legumes, Andrew Blake, Natalie Lauritsen, Department of Agriculture 18. Selecting the right variety for phase pasture systems, Keith Devenish, Department of Agriculture 19. Responses of alternative annual pasture and forage legumes to challenge with infectious subterranean clover mottle virus, John Fosu-Nyarko, Roger Jones, Lisa Smith, Mike Jones and Geoff Dwyer, State Agricultural Biotechnology Centre and Centre for Bioinformatics and Biological Computing, Murdoch University, Department of Agriculture, and Centre for Legumes in Mediterranean Agriculture SOIL AND LAND MANAGEMENT 20. Nutrition in 2002: Decisions to be made as a result of last season, Bill Bowden,Western Australia Department of Agriculture 21. Profitability of deep banding lime, Michael O\u27Connell, Chris Gazey and David Gartner, Department of Agriculture 22. Lime efficiency percentage…the new measure of lime effectiveness for Western Australia, Amanda Miller, Department of Agriculture 23. Boron – should we be worried about it, Richard W. BellA, K. FrostA, Mike WongBand Ross BrennanC ASchool of Environmental Science, Murdoch University, BCSIRO Land and Water, CDepartment of Agriculture 24. Impact of claying and other amelioration on paddock profit, N.J. Blake1, G. McConnell2, D. Patabendige1and N. Venn11Department of Agriculture, 2PlanFarm P/L 25. Raised bed farming in the 2001 growing season, Derk Bakker, Greg Hamilton, Dave Houlbrooke and Cliff Spann, Department of Agriculture 26. Economics of tramline farming systems, Paul Blackwell and Bindi Webb, Department of Agriculture, Stuart McAlpine, Liebe Group. 27. Relay planting from Tramlines to increase water use and productivity os summer crops, Dr Paul Blackwell, Department of Agriculture, Neil and Kim Diamond, Buntine. Liebe Group 28.Evidence-based zone management of paddock variability to improve profits and environmental outcomes, M.T.F. WongA, D. PatabendigeB, G. LyleA and K. WittwerA ACSIRO Land and Water, BDepartment of Agriculture 29. How much soil water is lost over summer in sandy soils? Perry Dolling1, Senthold Asseng2, Ian Fillery2, Phil Ward2and Michael Robertson3 1University of Western Australia/Department of Agriculture Western Australia/CSIRO, 2CSIRO Plant Industry 3CSIRO Sustainable Ecosystems, Indooroopilly, Queensland FARMER DECISION SUPPORT AND ADOPTION 30. Economic comparisons of farming systems for the medium rainfall northern sandplain, No 1, Caroline Peek and David Rogers, Department of Agriculture 31. Sensitivity analysis of farming systems for the medium rainfall northern sandplain No 2, Caroline Peek and David Rogers, Department of Agriculture 32. Transition analysis of farming systems in the medium rainfall northern sandplain. No 3, Caroline Peek and David Rogers, Department of Agriculture 33. Implementing on-farm quality assurance, Peter Portmann, Manager Research and Development, The Grain Pool of Western Australia 34. On-farm research – principles of the ‘Test As You Grow’ kit, Jeff Russell, Department of Agriculture 35. Broadscale wheat variety comparisons featuring Wyalkatchem, Jeff Russell, Department of Agriculture 36. GrainGuardÔ - A biosecurity plan for the Canola Industry,Greg Shea Department of Agriculture 37. Are Western Australian broadacre farms efficient? Ben Henderson, University of Western Australia, Ross Kingwell, Department of Agriculture and University of Western Australia DISEASE MODELLING WORKSHOP 38. WORKSHOP: Pest and disease forecasts for you! An interactive forum, Tresslyn Walmsley, Jean Galloway, Debbie Thackray, Moin Salam and Art Diggle, Centre for Legumes in Mediterranean Agriculture and Department of Agriculture 39. Blackspot spread: Disease models are based in reality (Workshop paper 1), JeanGalloway,Department of Agriculture 40. Blackspot spread: Scaling-up field data to simulate ‘Baker’s farm’ (Workshop paper 2), Moin U. Salam, Jean Galloway, Art J. Diggle and William J. MacLeod, Department of Agriculture, Western Australia 41. A decision support system for control of aphids and CMV in lupin crops (Workshop paper 3), Debbie Thackray, Jenny Hawkes and Roger Jones, Centre for Legumes in Mediterranean Agriculture and Department of Agricultur

Convalescent plasma in patients admitted to hospital with COVID-19 (RECOVERY): a randomised controlled, open-label, platform trial

Background: Many patients with COVID-19 have been treated with plasma containing anti-SARS-CoV-2 antibodies. We aimed to evaluate the safety and efficacy of convalescent plasma therapy in patients admitted to hospital with COVID-19. Methods: This randomised, controlled, open-label, platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]) is assessing several possible treatments in patients hospitalised with COVID-19 in the UK. The trial is underway at 177 NHS hospitals from across the UK. Eligible and consenting patients were randomly assigned (1:1) to receive either usual care alone (usual care group) or usual care plus high-titre convalescent plasma (convalescent plasma group). The primary outcome was 28-day mortality, analysed on an intention-to-treat basis. The trial is registered with ISRCTN, 50189673, and ClinicalTrials.gov, NCT04381936. Findings: Between May 28, 2020, and Jan 15, 2021, 11558 (71%) of 16287 patients enrolled in RECOVERY were eligible to receive convalescent plasma and were assigned to either the convalescent plasma group or the usual care group. There was no significant difference in 28-day mortality between the two groups: 1399 (24%) of 5795 patients in the convalescent plasma group and 1408 (24%) of 5763 patients in the usual care group died within 28 days (rate ratio 1·00, 95% CI 0·93–1·07; p=0·95). The 28-day mortality rate ratio was similar in all prespecified subgroups of patients, including in those patients without detectable SARS-CoV-2 antibodies at randomisation. Allocation to convalescent plasma had no significant effect on the proportion of patients discharged from hospital within 28 days (3832 [66%] patients in the convalescent plasma group vs 3822 [66%] patients in the usual care group; rate ratio 0·99, 95% CI 0·94–1·03; p=0·57). Among those not on invasive mechanical ventilation at randomisation, there was no significant difference in the proportion of patients meeting the composite endpoint of progression to invasive mechanical ventilation or death (1568 [29%] of 5493 patients in the convalescent plasma group vs 1568 [29%] of 5448 patients in the usual care group; rate ratio 0·99, 95% CI 0·93–1·05; p=0·79). Interpretation: In patients hospitalised with COVID-19, high-titre convalescent plasma did not improve survival or other prespecified clinical outcomes. Funding: UK Research and Innovation (Medical Research Council) and National Institute of Health Research

Effects of fluoxetine on functional outcomes after acute stroke (FOCUS): a pragmatic, double-blind, randomised, controlled trial

Background Results of small trials indicate that fluoxetine might improve functional outcomes after stroke. The FOCUS trial aimed to provide a precise estimate of these effects. Methods FOCUS was a pragmatic, multicentre, parallel group, double-blind, randomised, placebo-controlled trial done at 103 hospitals in the UK. Patients were eligible if they were aged 18 years or older, had a clinical stroke diagnosis, were enrolled and randomly assigned between 2 days and 15 days after onset, and had focal neurological deficits. Patients were randomly allocated fluoxetine 20 mg or matching placebo orally once daily for 6 months via a web-based system by use of a minimisation algorithm. The primary outcome was functional status, measured with the modified Rankin Scale (mRS), at 6 months. Patients, carers, health-care staff, and the trial team were masked to treatment allocation. Functional status was assessed at 6 months and 12 months after randomisation. Patients were analysed according to their treatment allocation. This trial is registered with the ISRCTN registry, number ISRCTN83290762. Findings Between Sept 10, 2012, and March 31, 2017, 3127 patients were recruited. 1564 patients were allocated fluoxetine and 1563 allocated placebo. mRS data at 6 months were available for 1553 (99·3%) patients in each treatment group. The distribution across mRS categories at 6 months was similar in the fluoxetine and placebo groups (common odds ratio adjusted for minimisation variables 0·951 [95% CI 0·839–1·079]; p=0·439). Patients allocated fluoxetine were less likely than those allocated placebo to develop new depression by 6 months (210 [13·43%] patients vs 269 [17·21%]; difference 3·78% [95% CI 1·26–6·30]; p=0·0033), but they had more bone fractures (45 [2·88%] vs 23 [1·47%]; difference 1·41% [95% CI 0·38–2·43]; p=0·0070). There were no significant differences in any other event at 6 or 12 months. Interpretation Fluoxetine 20 mg given daily for 6 months after acute stroke does not seem to improve functional outcomes. Although the treatment reduced the occurrence of depression, it increased the frequency of bone fractures. These results do not support the routine use of fluoxetine either for the prevention of post-stroke depression or to promote recovery of function. Funding UK Stroke Association and NIHR Health Technology Assessment Programme

Dimethyl fumarate in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Dimethyl fumarate (DMF) inhibits inflammasome-mediated inflammation and has been proposed as a treatment for patients hospitalised with COVID-19. This randomised, controlled, open-label platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing multiple treatments in patients hospitalised for COVID-19 (NCT04381936, ISRCTN50189673). In this assessment of DMF performed at 27 UK hospitals, adults were randomly allocated (1:1) to either usual standard of care alone or usual standard of care plus DMF. The primary outcome was clinical status on day 5 measured on a seven-point ordinal scale. Secondary outcomes were time to sustained improvement in clinical status, time to discharge, day 5 peripheral blood oxygenation, day 5 C-reactive protein, and improvement in day 10 clinical status. Between 2 March 2021 and 18 November 2021, 713 patients were enroled in the DMF evaluation, of whom 356 were randomly allocated to receive usual care plus DMF, and 357 to usual care alone. 95% of patients received corticosteroids as part of routine care. There was no evidence of a beneficial effect of DMF on clinical status at day 5 (common odds ratio of unfavourable outcome 1.12; 95% CI 0.86-1.47; p = 0.40). There was no significant effect of DMF on any secondary outcome

Dimethyl fumarate in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Dimethyl fumarate (DMF) inhibits inflammasome-mediated inflammation and has been proposed as a treatment for patients hospitalised with COVID-19. This randomised, controlled, open-label platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing multiple treatments in patients hospitalised for COVID-19 (NCT04381936, ISRCTN50189673). In this assessment of DMF performed at 27 UK hospitals, adults were randomly allocated (1:1) to either usual standard of care alone or usual standard of care plus DMF. The primary outcome was clinical status on day 5 measured on a seven-point ordinal scale. Secondary outcomes were time to sustained improvement in clinical status, time to discharge, day 5 peripheral blood oxygenation, day 5 C-reactive protein, and improvement in day 10 clinical status. Between 2 March 2021 and 18 November 2021, 713 patients were enroled in the DMF evaluation, of whom 356 were randomly allocated to receive usual care plus DMF, and 357 to usual care alone. 95% of patients received corticosteroids as part of routine care. There was no evidence of a beneficial effect of DMF on clinical status at day 5 (common odds ratio of unfavourable outcome 1.12; 95% CI 0.86-1.47; p = 0.40). There was no significant effect of DMF on any secondary outcome

Ribosomal Protein S6 Associates with Alphavirus Nonstructural Protein 2 and Mediates Expression from Alphavirus Messages

Although alphaviruses dramatically alter cellular function within hours of infection, interactions between alphaviruses and specific host cellular proteins are poorly understood. Although the alphavirus nonstructural protein 2 (nsP2) is an essential component of the viral replication complex, it also has critical auxiliary functions that determine the outcome of infection in the host. To gain a better understanding of nsP2 function, we sought to identify cellular proteins with which Venezuelan equine encephalitis virus nsP2 interacted. We demonstrate here that nsP2 associates with ribosomal protein S6 (RpS6) and that nsP2 is present in the ribosome-containing fractions of a polysome gradient, suggesting that nsP2 associates with RpS6 in the context of the whole ribosome. This result was noteworthy, since viral replicase proteins have seldom been described in direct association with components of the ribosome. The association of RpS6 with nsP2 was detected throughout the course of infection, and neither the synthesis of the viral structural proteins nor the presence of the other nonstructural proteins was required for RpS6 interaction with nsP2. nsP1 also was associated with RpS6, but other nonstructural proteins were not. RpS6 phosphorylation was dramatically diminished within hours after infection with alphaviruses. Furthermore, a reduction in the level of RpS6 protein expression led to diminished expression from alphavirus subgenomic messages, whereas no dramatic diminution in cellular translation was observed. Taken together, these data suggest that alphaviruses alter the ribosome during infection and that this alteration may contribute to differential translation of host and viral messages