Presented here is a magnetic hydrogel particle enabled workflow for capturing and concentrating SARS-CoV-2 from diagnostic remnant swab samples that significantly improves sequencing results using ...the Oxford Nanopore Technologies MinION sequencing platform. Our approach utilizes a novel affinity-based magnetic hydrogel particle, circumventing low input sample volumes and allowing for both rapid manual and automated high throughput workflows that are compatible with Nanopore sequencing. This approach enhances standard RNA extraction protocols, providing up to 40 × improvements in viral mapped reads, and improves sequencing coverage by 20-80% from lower titer diagnostic remnant samples. Furthermore, we demonstrate that this approach works for contrived influenza virus and respiratory syncytial virus samples, suggesting that it can be used to identify and improve sequencing results of multiple viruses in VTM samples. These methods can be performed manually or on a KingFisher automation platform.
Here we present a rapid and versatile method for capturing and concentrating SARS-CoV-2 from contrived transport medium and saliva samples using affinity-capture magnetic hydrogel particles. We ...demonstrate that the method concentrates virus from 1 mL samples prior to RNA extraction, substantially improving detection of virus using real-time RT-PCR across a range of viral titers (100-1,000,000 viral copies/mL) and enabling detection of virus using the 2019 nCoV CDC EUA Kit down to 100 viral copies/mL. This method is compatible with commercially available nucleic acid extraction kits (i.e., from Qiagen) and a simple heat and detergent method that extracts viral RNA directly off the particle, allowing a sample processing time of 10 min. We furthermore tested our method in transport medium diagnostic remnant samples that previously had been tested for SARS-CoV-2, showing that our method not only correctly identified all positive samples but also substantially improved detection of the virus in low viral load samples. The average improvement in cycle threshold value across all viral titers tested was 3.1. Finally, we illustrate that our method could potentially be used to enable pooled testing, as we observed considerable improvement in the detection of SARS-CoV-2 RNA from sample volumes of up to 10 mL.
The role of different project management techniques to implement projects successfully has been widely established in areas such as the planning and control of time, cost and quality. In spite of ...this the distinction between the project and project management is less than precise. This paper aims to identify the overlap between the definition of the project and project management and to discuss how the confusion between the two may affect their relationship. It identifies the different individuals involved on the project and project management, together with their objectives, expectations and influences. It demonstrates how a better appreciation of the distinction between the two will bring a higher possibility of project success.
The anticonvulsant phenytoin (5,5-diphenylhydantoin) provokes a skin rash in 5 to 10% of patients, which heralds the start of an idiosyncratic reaction that may result from covalent modification of ...normal self proteins by reactive drug metabolites. Phenytoin is metabolized by cytochrome P450 (P450) enzymes primarily to 5-(p-hydroxyphenyl-),5-phenylhydantoin (HPPH), which may be further metabolized to a catechol that spontaneously oxidizes to semiquinone and quinone species that covalently modify proteins. The aim of this study was to determine which P450s catalyze HPPH metabolism to the catechol, proposed to be the final enzymatic step in phenytoin bioactivation. Recombinant human P450s were coexpressed with NADPH-cytochrome P450 reductase in Escherichia coli. Novel bicistronic expression vectors were constructed for P450 2C19 and the three major variants of P450 2C9, i.e., 2C9*1, 2C9*2, and 2C9*3. HPPH metabolism and covalent adduct formation were assessed in parallel. P450 2C19 was the most effective catalyst of HPPH oxidation to the catechol metabolite and was also associated with the highest levels of covalent adduct formation. P450 3A4, 3A5, 3A7, 2C9*1, and 2C9*2 also catalyzed bioactivation of HPPH, but to a lesser extent. Fluorographic analysis showed that the major targets of adduct formation in bacterial membranes were the catalytic P450 forms, as suggested from experiments with human liver microsomes. These results suggest that P450 2C19 and other forms from the 2C and 3A subfamilies may be targets as well as catalysts of drug-protein adduct formation from phenytoin.
This participatory action research study was situated within a larger research study, which investigated the development of a peer-led Aboriginal parent support program in the remote setting of Halls ...Creek in Western Australia. The aim of this smaller research component was to explore the self-perceived role of home visiting peer support workers in this remote area. Participatory Action Research methodology was employed, within which eleven Action Learning Sets were used to facilitate participant collaboration (n = 8). All participant data was collected in focus groups during the Action Learning Sets where culturally appropriate home visiting parent support strategies were developed and examined in partnership with a child health nurse researcher. Data were analyzed using thematic analysis with three themes relating to the peer support workers' self-perceived role being identified: Helping parents be strong for their children; Strengthening culture; and Facilitating effective communication. These elements support peer support worker activities, helping parents to enhance their children's physical and psychosocial developmental environment. This research highlights the critical emerging role of peer support workers in home visiting family support in a remote area of Australia.
Joint Ventures, formed by organisations from two or more countries, have become a widespread form of strategic alliance in the world economy. They offer unique benefits of integrating the ...complimentary skills of different organisations cross culturally, combined with quicker market access, and satisfying the host nation’s technological gaps. This paper concentrates on the important features of Joint Venture formation in the former Soviet Union; exploring the negotiation of Joint Ventures, including sources of potential conflict. Changing economic policies within the former Soviet Union gave rise to particular concerns about their current management system. Some managerial issues arising in the former Soviet Union system are examined, with recommendations for methods that Venture partners can employ to overcome these problems. The paper is illustrated by a case from the development of an oil field in Western Kazakhstan.
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