Differential diagnostics of haemorrhagic fevers in resource poor environments.

This project centres on the proof of concept of a simple, rapid diagnostic platform for the differentiation of viral haemorrhagic fevers in resource poor environments. The benefits of this approach; 1. Simplicity, requiring only a fingerprick of blood. 2. Reduced time to detection, from blood to result in around 30 mins. 3. Suited to resource poor environments, removes the requirement for lab facilities or cold-chain.Portable and as such can be used in remote areas. 4. A molecular diagnostic assay, the system can differentiate and quantify a large number of viral targets and be sensitive enough to detect down to the 1000 viruses per ml level. 5. Reduced costs per test, removing the requirement for a lab while allowing high level multiplexing make the approach compare well with multiple immunological based tests while providing viral load information. 6. Reduced hazard risk in testing to the medical professional; assay process is easy to use even with users deploying bio-hazard ptotection and the assay is closed tube. During the recent Ebola outbreak BG Research (BGR) was tasked with adapting its technology for the direct detection of sepsis causing organisms to the rapid, closed tube detection of Ebola in remote areas. The approach migrated the proven CDC assay onto a novel instrument capable of detecting down to as low as 20,000 viruses per ml of blood - the same sensitivity as the laboratory based approach. That project adapted existing BGR instrumentation into a new technology demonstrator and the goals of this project are to develop a new prototype featuring a larger reaction volume such that the sensitivity can be improved by over 20 times - enough to potentially detect infection in convalescent patients and to monitor close contacts of infected patients not yet displaying symptoms. The main goals of the project are firstly to demonstrate a novel first step that concurrently renders the virus non-infectious while releasing the viral RNA - this involves rapidly freezing and thawing the blood sample in a buffer containing an RNA stabilising biocide. The whole process takes place in a single tube and during this project a large reaction vessel, capable of analysing 5 times more blood than the Ebola technology demonstrator is the 2nd goal. A prototype instrument, capable of in-field use and able to process multiple samples concurrently in a random access manner will be designed. Lastly, assays will be developed to demonstrate the system’s capability to differentiate large numbers of different viral targets concurrently in a multiplexed assay. This is a platform technology, but the final assay chosen here will be an RT-QPCR molecular differential assay for Ebola, Lassa, Marburg, Crimean-Congo fever and Rift valley fever to be tested on live virus at PHE Porton. The platform technology can be adapted to other blood-borne diseases such as Zika, Dengue, Chikungunya and Malaria dependent on regional requirements.