Antiretroviral Exposure within the Central Nervous System of Preclinical Models of HIV Infection

HIV reservoirs are major barriers to the eradication of this infection despite the widespread use of combination antiretroviral therapy. Antiretroviral drugs have restricted entry into the brain due to the inherent complexities of this tissue (e.g. uniquely structured blood-brain barrier, presence of efficient efflux pumps) which makes the brain an ideal potential anatomic reservoir for HIV replication. Cerebrospinal fluid (CSF) and plasma blood concentrations are often used to approximate drug concentrations within brain tissue; however, for reasons previously listed, drug concentrations in these compartments do not necessarily reflect those in the brain. The objective of this study was to evaluate the extent of penetration of select antiretroviral agents into the brain tissue of two preclinical models of HIV infection: rhesus macaques (n=6) and humanized mice (n=12). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to quantify efavirenz (EFV), emtricitabine (FTC), and tenofovir (TFV) in blood plasma, CSF, and brain tissue. EFV penetrated best into brain tissue in both animal models, and was up to 3.1-fold higher in macaques versus mice (p=0.026). FTC and TFV penetration into brain tissue was 1.3 to 25-fold higher macaques (p=0.004 and p=0.093, respectively). In infected macaques, the CSF-to-plasma ratios (CPRs) for TFV and FTC were 1.6 and 4-fold higher than the brain tissue-to-plasma ratio (TPR). CPRs for the remaining macaque samples were 1.3 to 61-fold lower than the TPRs, suggesting that more drug is getting into the brain tissue compared to the CSF. These results illustrate that CSF drug concentrations may provide underestimations of the penetration of antiretrovirals into brain tissue, and that HIV infection may result in lower brain tissue drug concentrations.Doctor of Pharmac

Epigenetic modifications and conserved, non-coding DNA play a role in regulation of type IV collagen gene expression

Type IV collagens are components of basement membranes throughout the body and are involved in maintenance of the structural integrity of tissues as well as cellular differentiation, growth, and adhesion. Members of this collagen family are uniquely arranged in pairs in a head-to-head orientation and share a proximal promoter region. The COL4A5-COL4A6 gene pair is involved in numerous human diseases and cancer metastasis. For these reasons, defining the mechanisms that regulate collagen gene expression is of specific interest. To study type IV collagens, an in vitro model system was characterized. Comparative genomics was utilized to identify conserved, non-coding DNA in COL4A5 and COL4A6. These sequences were transfected into cell lines differing in type IV collagen expression and tested for the ability to regulate transcription of a reporter gene. Each cell line was also treated with the epigenetic modifying agents, 5-Aza and TSA. The effects on type IV collagen expression were determined. The COL4A5-COL4A6 promoter region was extensively characterized using ChIP analysis; antibodies against RNAPII, acetylated histone H3, and H3K9me2 were used. Additionally, bisulfite sequencing was carried out on each cell line to determine the methylation status of CpG dinucleotides in the promoter. Cell lines differing in expression of COL4A5 and COL4A6 were identified: 1) SCC-25 keratinocytes and HEK-293 cells transcribed both COL4A5 and COL4A6, 2) HT-1080 cells selectively activated COL4A5, and 3) SK-N-SH neuroblastoma cells did not express either gene. In SK-N-SH cells, histone modifications were shown to facilitate formation of condensed chromatin to prevent transcription initiation; repression was independent of DNA methylation. Activation of COL4A5 and COL4A6 in SCC-25 and HEK-293 cells involved acetylation of histones, although differences between the two cell types were seen. In addition, conserved, non-coding sequences were shown to affect transcription of a reporter gene; these sequences may be interacting with the transcription machinery to modulate collagen expression. Finally, repression of COL4A6 in HT-1080 cells appeared to be mediated through DNA methylation of the promoter; selective activation of COL4A5 may involve conserved, non-coding DNA. In summary, epigenetic modifications as well as conserved sequences are intimately involved in regulation of type IV collagen gene expression

SNX27–Retromer directly binds ESCPE-1 to transfer cargo proteins during endosomal recycling

Coat complexes coordinate cargo recognition through cargo adaptors with biogenesis of transport carriers during integral membrane protein trafficking. Here, we combine biochemical, structural, and cellular analyses to establish the mechanistic basis through which SNX27-Retromer, a major endosomal cargo adaptor, couples to the membrane remodeling endosomal SNX-BAR sorting complex for promoting exit 1 (ESCPE-1). In showing that the SNX27 FERM (4.1/ezrin/radixin/moesin) domain directly binds acidic-Asp-Leu-Phe (aDLF) motifs in the SNX1/SNX2 subunits of ESCPE-1, we propose a handover model where SNX27-Retromer captured cargo proteins are transferred into ESCPE-1 transport carriers to promote endosome-to-plasma membrane recycling. By revealing that assembly of the SNX27:Retromer:ESCPE-1 coat evolved in a stepwise manner during early metazoan evolution, likely reflecting the increasing complexity of endosome-to-plasma membrane recycling from the ancestral opisthokont to modern animals, we provide further evidence of the functional diversification of yeast pentameric Retromer in the recycling of hundreds of integral membrane proteins in metazoans

Efficacy of FFP3 respirators for prevention of SARS-CoV-2 infection in healthcare workers.

Funder: Addenbrooke's Charitable Trust, Cambridge University Hospitals; FundRef: http://dx.doi.org/10.13039/501100002927Funder: NIHR Cambridge Biomedical Research CentreBackground: Respiratory protective equipment recommended in the UK for healthcare workers (HCWs) caring for patients with COVID-19 comprises a fluid-resistant surgical mask (FRSM), except in the context of aerosol generating procedures (AGPs). We previously demonstrated frequent pauci- and asymptomatic severe acute respiratory syndrome coronavirus 2 infection HCWs during the first wave of the COVID-19 pandemic in the UK, using a comprehensive PCR-based HCW screening programme (Rivett et al., 2020; Jones et al., 2020). Methods: Here, we use observational data and mathematical modelling to analyse infection rates amongst HCWs working on 'red' (coronavirus disease 2019, COVID-19) and 'green' (non-COVID-19) wards during the second wave of the pandemic, before and after the substitution of filtering face piece 3 (FFP3) respirators for FRSMs. Results: Whilst using FRSMs, HCWs working on red wards faced an approximately 31-fold (and at least fivefold) increased risk of direct, ward-based infection. Conversely, after changing to FFP3 respirators, this risk was significantly reduced (52-100% protection). Conclusions: FFP3 respirators may therefore provide more effective protection than FRSMs for HCWs caring for patients with COVID-19, whether or not AGPs are undertaken. Funding: Wellcome Trust, Medical Research Council, Addenbrooke's Charitable Trust, NIHR Cambridge Biomedical Research Centre, NHS Blood and Transfusion, UKRI

Evaluation of three parasite lactate dehydrogenase-based rapid diagnostic tests for the diagnosis of falciparum and vivax malaria

BACKGROUND: In areas where non-falciparum malaria is common rapid diagnostic tests (RDTs) capable of distinguishing malaria species reliably are needed. Such tests are often based on the detection of parasite lactate dehydrogenase (pLDH). METHODS: In Dawei, southern Myanmar, three pLDH based RDTs (CareStart Malaria pLDH (Pan), CareStart Malaria pLDH (Pan, Pf) and OptiMAL-IT)were evaluated in patients presenting with clinically suspected malaria. Each RDT was read independently by two readers. A subset of patients with microscopically confirmed malaria had their RDTs repeated on days 2, 7 and then weekly until negative. At the end of the study, samples of study batches were sent for heat stability testing. RESULTS: Between August and November 2007, 1004 patients aged between 1 and 93 years were enrolled in the study. Slide microscopy (the reference standard) diagnosed 213 Plasmodium vivax (Pv) monoinfections, 98 Plasmodium falciparum (Pf) mono-infections and no malaria in 650 cases. The sensitivities (sens) and specificities (spec), of the RDTs for the detection of malaria were- CareStart Malaria pLDH (Pan) test: sens 89.1% [CI95 84.2-92.6], spec 97.6% [CI95 96.5-98.4]. OptiMal-IT: Pf+/- other species detection: sens 95.2% [CI95 87.5-98.2], spec 94.7% [CI95 93.3-95.8]; non-Pf detection alone: sens 89.6% [CI95 83.6-93.6], spec 96.5% [CI95 94.8-97.7]. CareStart Malaria pLDH (Pan, Pf): Pf+/- other species: sens 93.5% [CI95 85.4-97.3], spec 97.4% [95.9-98.3]; non-Pf: sens 78.5% [CI95 71.1-84.4], spec 97.8% [CI95 96.3-98.7]. Inter-observer agreement was excellent for all tests (kappa > 0.9). The median time for the RDTs to become negative was two days for the CareStart Malaria tests and seven days for OptiMAL-IT. Tests were heat stable up to 90 days except for OptiMAL-IT (Pf specific pLDH stable to day 20 at 35 degrees C). CONCLUSION: None of the pLDH-based RDTs evaluated was able to detect non-falciparum malaria with high sensitivity, particularly at low parasitaemias. OptiMAL-IT performed best overall and would perform best in an area of high malaria prevalence among screened fever cases. However, heat stability was unacceptable and the number of steps to perform this test is a significant drawback in the field. A reliable, heat-stable, highly sensitive RDT, capable of diagnosing all Plasmodium species has yet to be identified

Staged induction of HIV-1 glycan–dependent broadly neutralizing antibodies

A preventive HIV-1 vaccine should induce HIV-1–specific broadly neutralizing antibodies (bnAbs). However, bnAbs generally require high levels of somatic hypermutation (SHM) to acquire breadth, and current vaccine strategies have not been successful in inducing bnAbs. Because bnAbs directed against a glycosylated site adjacent to the third variable loop (V3) of the HIV-1 envelope protein require limited SHM, the V3-glycan epitope is an attractive vaccine target. By studying the cooperation among multiple V3-glycan B cell lineages and their coevolution with autologous virus throughout 5 years of infection, we identify key events in the ontogeny of a V3-glycan bnAb. Two autologous neutralizing antibody lineages selected for virus escape mutations and consequently allowed initiation and affinity maturation of a V3-glycan bnAb lineage. The nucleotide substitution required to initiate the bnAb lineage occurred at a low-probability site for activation-induced cytidine deaminase activity. Cooperation of B cell lineages and an improbable mutation critical for bnAb activity defined the necessary events leading to breadth in this V3-glycan bnAb lineage. These findings may, in part, explain why initiation of V3-glycan bnAbs is rare, and suggest an immunization strategy for inducing similar V3-glycan bnAbs

Crop Updates 2001 - Cereals

This session covers forty two papers from different authors: PLENARY 1. Planning your cropping program in season 2001, Dr Ross Kingwell, Agriculture Western Australia and University of Western Australia WORKSHOP 2. Can we produce high yields without high inputs? Wal Anderson, Centre for Cropping Systems, Agriculture Western Australia VARIETIES 3. Local and interstate wheat variety performance and $ return to WA growers, Eddy Pol, Peter Burgess and Ashley Bacon, Agritech Crop Research CROP ESTABLISHMENT 4 Soil management of waterlogged soils, D.M. Bakker, G.J. Hamilton, D. Houlbrooke and C. Spann, Agriculture Western Australia 5. Effect of soil amelioration on wheat yield in a very dry season, M.A Hamza and W.K. Anderson, Agriculture Western Australia 6. Fuzzy tramlines for more yield and less weed, Paul Blackwell1 and Maurice Black2 1Agriculture Western Australia, 2Harbour Lights Estate, Geraldton 7. Tramline farming for dollar benefits, Paul Blackwell, Agriculture Western Australia NUTRITION 8. Soil immobile nutrients for no-till crops, M.D.A. Bolland1, R.F. Brennan1,and W.L. Crabtree2, 1Agriculture Western Australia, 2Western Australian No-Tillage Farmers Association 9. Burn stubble windrows: to diagnose soil fertility problems, Bill Bowden, Chris Gazey and Ross Brennan, Agriculture Western Australia 10. Calcium: magnesium ratios; are they important? Bill Bowden1, Rochelle Strahan2, Bob Gilkes2 and Zed Rengel2 1Agriculture Western Australia, 2Department of Soil Science and Plant Nutrition, UWA 11. Responses to late foliar applications of Flexi-N, Stephen Loss, Tim O’Dea, Patrick Gethin, Ryan Guthrie, Lisa Leaver, CSBP futurefarm 12. A comparison of Flexi-N placements, Stephen Loss, Tim O’Dea, Patrick Gethin, Ryan Guthrie, Lisa Leaver, CSBP futurefarm 13. What is the best way to apply potassium? Stephen Loss, Tim O’Dea, Patrick Gethin, Ryan Guthrie, CSBP futurefarm 14. Claying affects potassium nutrition in barley, Stephen Loss, David Phelps, Tim O’Dea, Patrick Gethin, Ryan Guthrie, Lisa Leaver, CSBP futurefarm 15. Nitrogen and potassium improve oaten hay quality, Stephen Loss, Tim O’Dea, Patrick Gethin, Ryan Guthrie, Lisa Leaver, CSBP futurefarm AGRONOMY 16. Agronomic responses of new wheat varieties in the northern wheatbelt, Darshan Sharma and Wal Anderson, Agriculture Western Australia 17. Wheat agronomy research on the south coast, Mohammad Amjad and Wal Anderson, Agriculture Western Australia 18. Influence of sowing date on wheat yield and quality in the south coast environment, Mohammad Amjadand Wal Anderson, Agriculture Western Australia 19. More profit from durum, Md.Shahajahan Miyan and Wal Anderson, Agriculture Western Australia 20. Enhancing recommendations of flowering and yield in wheat, JamesFisher1, Senthold Asseng2, Bill Bowden1 and Michael Robertson3 ,1AgricultureWestern Australia, 2CSIRO Plant Industry, 3CSIRO Sustainable Ecosystems 21. When and where to grow oats, Glenn McDonald, Agriculture Western Australia 22. Managing Gaidner barley for quality, Kevin Young and Blakely Paynter, Agriculture Western Australia PESTS AND DISEASES 23. Strategies for leaf disease management in wheat, Jatinderpal Bhathal1, Cameron Weeks2, Kith Jayasena1 and Robert Loughman1 ,1Agriculture Western Australia. 2Mingenew-Irwin Group Inc 24. Strategies for leaf disease management in malting barley, K. Jayasena1, Q. Knight2 and R. Loughman1, 1Agriculture Western Australia, 2IAMA Agribusiness 25. Cereal disease diagnostics, Dominie Wright and Nichole Burges, Agriculture Western Australia 26. The big rust: Did you get your money back!! Peter Burgess, Agritech Crop Research 27. Jockey – winning the race against disease in wheat, Lisa-Jane Blacklow, Rob Hulme and Rob Giffith, Aventis CropScience 28. Distribution and incidence of aphids and barley yellow dwarf virus in over-summering grasses in WA wheatbelt, Jenny Hawkes and Roger Jones, CLIMA and Agriculture Western Australia 29. Further developments in forecasting aphid and virus risk in cereals, Debbie Thackray, Jenny Hawkes and Roger Jones, Agriculture Western Australia and Centre for Legumes in Mediterranean Agriculture 30. Effect of root lesion nematodes on wheat yields in Western Australia, S. B. Sharma, S. Kelly and R. Loughman, Crop Improvement Institute, Agriculture Western Australia 31. Rotational crops and varieties for management of root lesion nematodes in Western Australia, S.B. Sharma, S. Kelly and R. Loughman, Crop Improvement Institute, Agriculture Western Australia WEEDS 32. Phenoxy herbicide tolerance of wheat, Peter Newman and Dave Nicholson, Agriculture Western Australia 33. Tolerance of wheat to phenoxy herbicides,Harmohinder S. Dhammu, Terry Piper and Mario F. D\u27Antuono, Agriculture Western Australia 34. Herbicide tolerance of durum wheats, Harmohinder S. Dhammu, Terry Piper and David Nicholson, Agriculture Western Australia 35. Herbicide tolerance of new wheats, Harmohinder S. Dhammu, Terry Piper and David F. Nicholson, Agriculture Western Australia BREEDING 36. Towards molecular breeding of barley: construction of a molecular genetic map, Mehmet Cakir1, Nick Galwey1, David Poulsen2, Garry Ablett3, Reg Lance4, Rob Potter5 and Peter Langridge6,1Plant Sciences, Faculty of Agriculture, UWA, 2Queensland Department of Primary Industries, Qld, 3Centre for Plant Conservation Genetics Southern Cross University, Lismore NSW, 5SABC Murdoch University, WA, 6Department of Plant Science University of Adelaide, Glen Osmond SA 37. Toward molecular breeding of barley: Identifying markers linked to genes for quantitative traits, Mehmet Cakir1, Nick Galwey1, David Poulsen2, Reg Lance3, Garry Ablett4, Greg Platz2, Joe Panozzo5, Barbara Read6, David Moody5, Andy Barr7 and Peter Langridge7 , 1Plant Sciences, Faculty of Agriculture, UWA, 2Queensland Department of Primary Industries, Warwick, QLD,3Agriculture Western Australia, 4Centre for Plant Conservation Genetics, Southern Cross University, Lismore NSW, 5VIDA Private Bag 260, Horsham VIC, 6NSW Dept. of Agriculture, Wagga Wagga NSW, 7Department of Plant Science, University of Adelaide, Glen Osmond SA 38. Can we improve grain yield by breeding for greater early vigour in wheat? Tina Botwright1, Tony Condon1, Robin Wilson2 and Iain Barclay2, 1CSIRO Plant Industry, 2Agriculture Western Australia MARKETING AND QUALITY 39. The Crop Improvement Royalty, Howard Carr, Agriculture Western Australia 40. GrainGuardÔ - The development of a protection plan for the wheat industry, Greg Shea, Agriculture Western Australia CLIMATE 41. Rainfall – what happened in 2000 and the prospects for 2001, Ian Foster, Agriculture Western Australia 42. Software for climate management issues, David Tennant,Agriculture Western Australia CONTRIBUTING AUTHOR CONTACT DETAIL