Determining how atmospheric carbon dioxide concentrations have changed during the history of the Earth

The reconstruction of ancient atmospheric carbon dioxide concentrations is essential to understanding the history of the Earth and life. It is also an important guide to identifying the sensitivity of the Earth system to this greenhouse gas and, therefore, constraining its future impact on climate. However, determining the concentration of CO2 in ancient atmospheres is a challenging endeavour requiring the application of state-of-the-art analytical chemistry to geological materials, underpinned by an understanding of photosynthesis and biochemistry. It is truly an interdisciplinary challenge

Cell therapy-processing economics: small-scale microfactories as a stepping stone toward large-scale macrofactories

Aim: Manufacturing methods for cell-based therapies differ markedly from those established for noncellular pharmaceuticals and biologics. Attempts to ‘shoehorn’ these into existing frameworks have yielded poor outcomes. Some excellent clinical results have been realized, yet emergence of a ‘blockbuster’ cell-based therapy has so far proved elusive. Materials & methods: The pressure to provide these innovative therapies, even at a smaller scale, remains. In this process, economics research paper, we utilize cell expansion research data combined with operational cost modeling in a case study to demonstrate the alternative ways in which a novel mesenchymal stem cell-based therapy could be provided at small scale. Results & Conclusions: This research outlines the feasibility of cell microfactories but highlighted that there is a strong pressure to automate processes and split the quality control cost-burden over larger production batches. The study explores one potential paradigm of cell-based therapy provisioning as a potential exemplar on which to base manufacturing strategy

Basal rot of narcissus : understanding pathogenicity in fusarium oxysporum f. sp. narcissi

Fusarium oxysporum is a globally distributed soilborne fungal pathogen causing root rots, bulb rots, crown rots and vascular wilts on a range of horticultural plants. Pathogenic F. oxysporum isolates are highly host specific and are classified as formae speciales. Narcissus is an important ornamental crop and both the quality and yield of flowers and bulbs can be severely affected by a basal rot caused by F. oxysporum f. sp. narcissi (FON); 154 Fusarium isolates were obtained from different locations and Narcissus cultivars in the United Kingdom, representing a valuable resource. A subset of 30 F. oxysporum isolates were all found to be pathogenic and were therefore identified as FON. Molecular characterisation of isolates through sequencing of three housekeeping genes, suggested a monophyletic origin with little divergence. PCR detection of 14 Secreted in Xylem (SIX) genes, previously shown to be associated with pathogenicity in other F. oxysporum f. spp., revealed different complements of SIX7, SIX9, SIX10, SIX12 and SIX13 within FON isolates which may suggest a race structure. SIX gene sequences were unique to FON and SIX10 was present in all isolates, allowing for molecular identification of FON for the first time. The genome of a highly pathogenic isolate was sequenced and lineage specific (LS) regions identified which harboured putative effectors including the SIX genes. Real-time RT-PCR, showed that SIX genes and selected putative effectors were expressed in planta with many significantly upregulated during infection. This is the first study to characterise molecular variation in FON and provide an analysis of the FON genome. Identification of expressed genes potentially associated with virulence provides the basis for future functional studies and new targets for molecular diagnostics

Early Sowing and Irrigating of Rape Crops in Cool Temperate Environments Boosts Forage Yield Potential

Forage rape is commonly grown on Tasmanian dairy farms to provide feed during periods of low pasture growth and/or low nutritive value. In these regions, forage crops are generally sown between early spring and late autumn and are commonly rainfed, though farmers may apply a single irrigation at sowing. While the effect of water availability on forage yield has been well characterised for regions in north-western Tasmania (Neilsen 2005), there is a dearth of information for other regions in Tasmania. Hence, the objectives of this study were to identify optimal sowing times for, and determine the extent to which a single irrigation at sowing influences productivity of, rainfed brassica crops across the dairy regions of Tasmania

Impact of Grazing on the Silage Yield of Forage Oat Crops

Winter forage oat crops are increasing in acreage on Tasmanian dairy farms. Such crops can be grazed multiple times during vegetative development in winter then later harvested for silage at the booting or soft dough growth stages in spring. Although effects of grazing on grain yield of winter cereal crops have been well characterised (Harrison et al. 2011), little research has been conducted on the influence of grazing on the forage and silage yield potential of oats grown in Tasmania. We aimed to explore the influence of grazing management on the yields of forage for grazing and ensiling of oat crops using a biophysical crop model

Magnetic targeting of stem cell therapies

Stem cells may offer solutions for many health issues facing the world’s population. Early Biotech-led approaches are supporting novel mesenchymal stem cell (MSC) therapies through biomedical trials. However, their potential benefits are currently curtailed by challenges linked to high cell dose requirements which pose availability and manufacturing challenges, combined to suboptimal delivery methodologies. Whilst systemic delivery may be suitable for many pharmaceuticals, more complex and selective treatments such as emerging cell therapies require smarter targeting strategies on safety and cost/benefit grounds. Several groups are developing targeting strategies to guide stem cells to specific locations and hold them in situ whilst performing a repair. The targeting approach presented here uses superparamagnetic iron oxide microparticles (MPs) loaded within stem cells to facilitate control of the cells using magnets. Magnetic resonance imaging (MRI) can be used to monitor the loaded cells’ contribution to the repair process. Questions remain around MP safety and effects on both delivered cell therapies and the receiving patient. Presented data demonstrates labelling of MSC populations with a commercially available MP called SiMAG in two sizes (500 nm and 1000 nm). Particles were assessed for characteristics which influence their suitability for labelling and were found to have a non-uniform variable structure and size. Labelling was found to be both rapid and effective with low 10 µg/Fe/mL labelling doses able to distinguish cell populations by flow cytometry. Super-resolution microscopy, fluorescent microscopy and transmission electron microscopy were used to determine the location of particles within the cell and were noted to accumulate around the nucleus in large vesicles. Uptake into the cell was found to be influenced by serum with 10% serum resulting in a 75% drop in relative uptake over a 24 hour period. Potential sharing of MP between MSCs was investigated both qualitatively with fluorescent microscopy and quantitatively in a MSC co-culture experiment. No statistically significant sharing of MPs between MSCs could be seen to be taking place between populations. The fate of MPs within MSCs was investigated using pH nanosensors to interrogate the internal cell pH. A novel flow cytometry assay using pH nanosensors and MPs was used to examine the internal pH of large populations of cells. This yielded results which suggest a pH decrease over 4 days from pH 5.5 to 4.7 followed an increase to 5.4 by day 6. This effect is suggested to be caused by a complex pH mediated degradation of MPs followed by increase in pH due to the degraded iron overloading the cell. This degradation was carried out in simulated lysosomal conditions and found to act in a similar way. Macrophages were also used to test degradation and again they were found to reduce the fluorecense of the MPs rapidly over 7 days. The ability of MSCs to tolerate MPs without impacting cell health was probed with a range of assays. The metabolic assay Presto blue demonstrated doses of 10 µg/Fe/mL did not impact the metabolic status of the cell. This was tested with other potential surface chemistries of the same particle design and these were also found to be well tolerated. Membrane intergrity was assessed with flow cytometry for both 500 and 1000 nm SiMAG and was found to have no damaging effects present at 10 µg/Fe/mL. SiMAG 1000 nm was found to have no membrane compromising effect all the way up to 100 µg/Fe/mL. Cell identity was assessed with common MSC markers established by the Dominici position paper and no change to expression was found to occur even with repeated, high dose long-term (14 day) labelling strategies. As particles accumulate round the nucleus, deleterious effects of MP on DNA were tested using the comet assay and visual inspection with no statistically significant increase up to the maximum tested of 100 µg/Fe/mL. Similarly, no effect on cell cycle status was noticed for populations of MSCs. The retention of cell “function” was tested not only following labelling, but following hypothermic storage of cells to simulate shipping to a clinical setting. This was carried out for a range of clinically relevant cell types including mesenchymal stem cells (MSCs), cardiomyocytes (CaM) and ReNeuron neural stem cells (ReN). MSCs were found to freely differentiate to tri-lineage osteogenic, chondrogenic and adipogenic lineages. CXCR4 expression was measured as a marker of MSCs ability to home in on damage and was found to be raised in response to MP presence. CaM were found to resume beating both after hypothermic storage, as well as at high (1000 µg/Fe/mL SiMAG doses). ReN cells were found to be more sensitive to SiMAG with only 10 µg/Fe/mL doses tolerated although successful neural differentiation was still possible. The ability to culture MSCs and label them in a scalable manufacturing scenario was also examined and found to be possible. SiMAG was demonstrated to be a suitable labelling agent both for imaging as well as magnetic manipulation. Precise magnetic manipulation of labelled cells was demonstrated both as an individual cell and as a cell population moving through a simulated tissue gel. Entrapment of labelled cells from a simulated circulatory system was also shown to be possible with close to 100% of cells recruited in the first pass. The fluorophores on SiMAG were not strong enough to be visualised on their own and quantum dots were used to demonstrate successful retention of labelled MSCs in an ex vivo rat model. MRI was however shown to be a suitable method for assessing the location of labelled cell populations at even low cell concentrations ~1x106 and low SiMAG doses of 5 µf/Fe/mL. In conclusion, this study indicates that MPs represent a cell-safe and effective potential tool to better target cells. The key benefit of this cell labelling technology is in the high degree of understanding over the entire labelling process from entry through to degradation. In addition, this labelling technology has been shown to be cell-safe in a large number of cells both for physical health and basic function. Whilst this should be further explored for further, more specific applications, it makes a compelling case for SiMAG as a multi-functional tool for cell manipulation and tracking

Centralised versus decentralised manufacturing and the delivery of healthcare products: A United Kingdom exemplar

Background. The cell and gene therapy (CGT) field is at a critical juncture. Clinical successes have underpinned the requirement for developing manufacturing capacity suited to patient-specific therapies that can satisfy the eventual demand post-launch. Decentralised or ‘redistributed’ manufacturing divides manufacturing capacity across geographic regions, promising local, responsive manufacturing, customised to the end user, and is an attractive solution to overcome challenges facing the CGT manufacturing chain. Methods. A study was undertaken building on previous, so far unpublished, semistructured interviews with key opinion leaders in advanced therapy research, manufacturing and clinical practice.The qualitative findings were applied to construct a cost of goods model that permitted the cost impact of regional siting to be combined with variable and fixed costs of manufacture of a mesenchymal stromal cell product. Results. Using the United Kingdom as an exemplar, cost disparities between regions were examined. Per patient dose costs of ~£1,800 per 75,000,000 cells were observed. Financial savings from situating the facility outside of London allow 25–41 additional staff or 24–35 extra manufacturing vessels to be employed. Decentralised quality control to mitigate site-to-site variation was examined. Partial decentralisation of quality control was observed to be financially possible and an attractive option for facilitating release ‘at risk’. Discussion. There are important challenges that obstruct the easy adoption of decentralised manufacturing that have the potential to undermine the market success of otherwise promising products. By using the United Kingdom as an exemplar, the modelled data provide a framework to inform similar regional policy considerations across other global territories

Systematic reviews: guidance relevant for studies of older people

Systematic reviews and meta-analyses are increasingly common. This article aims to provide guidance for people conducting systematic reviews relevant to the healthcare of older people. An awareness of these issues will also help people reading systematic reviews to determine whether the results will influence their clinical practice. It is essential that systematic reviews are performed by a team which includes the required technical and clinical expertise. Those performing reviews for the first time should ensure they have appropriate training and support. They must be planned and performed in a transparent and methodologically robust way: guidelines are available. The protocol should be written—and if possible published—before starting the review. Geriatricians will be interested in a table of baseline characteristics, which will help to determine if the studied samples or populations are similar to their patients. Reviews of studies of older people should consider how they will manage issues such as different age cut-offs; non-specific presentations; multiple predictors and outcomes; potential biases and confounders. Systematic reviews and meta-analyses may provide evidence to improve older people's care, or determine where new evidence is required. Newer methodologies, such as meta-analyses of individual level data, network meta-analyses and umbrella reviews, and realist synthesis, may improve the reliability and clinical utility of systematic reviews

Identification of pathogenicity-related genes in Fusarium oxysporum f. sp. cepae

Pathogenic isolates of Fusarium oxysporum, distinguished as formae speciales (f. spp.) based on their host specificity, cause crown rots, root rots and vascular wilts on many important crops worldwide. F. oxysporum f. sp. cepae (FOC) is particularly problematic to onion growers worldwide and is increasing in prevalence in the UK. We characterised 31 F. oxysporum isolates collected from UK onions using pathogenicity tests, sequencing of housekeeping genes and identification of effectors. In onion seedling and bulb tests, 21 isolates were pathogenic while 10 were non-pathogenic. Molecular characterisation of these isolates, and 21 additional isolates comprising other f. spp. and different Fusarium species, was carried out by sequencing three housekeeping genes. A concatenated tree separated the F. oxysporum isolates into six clades but did not distinguish pathogenic and non-pathogenic isolates. Ten putative effectors were identified within FOC, including seven Secreted in Xylem (SIX) genes first reported in F. oxysporum f. sp. lycopersici. Two highly homologous proteins with signal peptides and RxLR motifs (CRX1/CRX2) and a gene with no previously characterised domains (C5) were also identified. The presence/absence of nine of these genes was strongly related to pathogenicity against onion and all were shown to be expressed in planta. Different SIX gene complements were identified in other f. spp. but none were identified in three other Fusarium species from onion. Although the FOC SIX genes had a high level of homology with other f. spp. there were clear differences in sequence which were unique to FOC while CRX1 and C5 genes appear to be largely FOC specific. This article is protected by copyright. All rights reserved