Over the course of the project, we were assisted by Gurpreet, a Nuffield Bursary Student from Whitley Academy in Coventry currently entering her 2nd year of sixth form. Through joining us she hoped to further her knowledge of biology and chemistry enabling her to make a more well-informed decision for University applications. In addition to teaching her practical skills in the lab, we also educated her on synthetic biology topics not covered in her current academic syllabus.

Gurpreet was very helpful in providing an alternative perspective as someone from outside the academic sphere with which to share ideas. By identifying the more complex concepts that were that were most difficult to explain, she helped us prepare for attending public events such as open days. Gurpreet expressed a keen interest in the different methods that could be applied when constructing our sensor in order to make it most suitable for use by a wide demographic. By encouraging us to consider the different environments in which our device could be applied, and further involving herself by researching potential solutions, she influenced the development of the physical composition of our device and method of delivery.

We were very fortunate to speak to David Lloyd, CEO of FredSense Technologies - a leading biosensor company based in Canada. He was able to advise our team with regards to the business side of our project. On his advice, we decided that our sensor should not cost more than 0.50 USD per sensor, in order to successfully target the developing world. When estimating manufacturing costs, David recommended considering pre-treatment requirements, such as purification of the RNA and the freeze-drying process, as well as any hardware needed. His general guide figure of the overall cost of production was less than 30% of the resale price. He also proposed additional applications for our sensors, including testing mine runoff and domestic wells in developed countries.

David identified numerous issues that FredSense determined may affect the success of biosensors. Illiteracy and poor governmental structure may prevent success in developing countries, while lack of available treatment may render identifying health hazards potentially pointless. To tackle these problems, the team proposed a colorimetric sensing system that could be supplied to charities and governmental aid associations. However, he did warn that colour is subjective, and so for application within more developed areas, such as supplying environmental consultancies, we determined that a more quantitative measurement would be more helpful. We also considered further developing our product by combining the sensor with a water purification treatment.

After more detailed discussion, the team gained a greater insight into the potential interference resulting from other metal ions present in the sensing solution – potassium for example, interferes with the detection of lead. FredSense found they had to take preventative measures to reduce the effects of arsenic on antimony detection in particular, an interaction which we had not previously considered. Following this, we decided to combat any potential interference issues we may have by treating sample water with 18-crown-6 ether. This reagent specifically binds and masks potassium and sodium ions in solution, preventing their interaction with our sensor and reducing the chance of false positives.

This year, two members of the Warwick iGEM team attended the annual Lyme Disease Association conference was held at Murray Edwards College, Cambridge University. The theme of the conference was ‘Moving Forward’, and focused on the progress that Lyme disease research has made. The first speaker was Dr Norma O’Flynn from the National Guideline Centre, elaborating upon the procedure applied by the National Institute for Health and Care Excellence, when determining management processes to improve health and social care. She discussed the different factors considered before a policy can be issued, and how this procedure is currently being applied to Lyme disease.

Dr Sandra Pearson, Lyme Disease Action, tackled the issue of Borrelia persistence, followed by Dr Monica Embers, from the Tulane National Primate Research Centre, who had modelled the changes in immune response over the course of infection and post treatment using rhesus macaques. Dr Embers also discussed her research into the challenges of diagnosing and curing Lyme disease. She reviewed the current testing methods for diagnosis – with early serology tests including IFA, ELISA, western blots and PCR, and later stage diagnostics testing for antigens/antibodies in CSF, urine and the blood. Dr Embers identified several issues with the current system - these tests are not specific or sensitive enough to diagnose patients at all phases of the disease, antigenic variability may prevent antibody recognition and western blots have a degree of inaccuracy when detecting the antibodies present. She proposed a more sensitive analysis technique that her team had been developing – a 5-antigen Bioplex assay.

During the afternoon session, Dr Jinyu Shan presented the University of Leicester's alternative diagnosis method – engineering bacteriophages to effectively kill Borrelia by overexpression of holins and endolysins (phage-encoded enzymes). According to Dr Shan, the pilot study indicated the novel diagnostic test had a 75% senstivity and 100% specificity on serum samples as small as 400 µl, as well as being able to distinguish between Lyme and relapsing fever Borrelia strains.

Being able to interact with research specialists, GP’s and actual Lyme disease sufferers was a fantastic opportunity to gauge first-hand the opinions of different demographics in response to our project. We spoke to several researchers investigating epidemiology, novel targets for diagnosis, and new treatments, as well as several patients who had recently finished their course of antibiotics following diagnosis. We were very fortunate to speak to Dr Tim Brooks – the current director of Public Health England’s Rare and Imported Pathogens Laboratory (RIPL). The RIPL act as the governmental body for acute diagnostic facilities for a wide range of diseases in the UK, including zoonotic vector-borne diseases such as Lyme disease. After discussing the advantages and limitations of the current serology and PCR diagnostic tests, we proposed our paper-based sensor and explained the testing procedure. He acknowledged the usefulness of a more robust, faster detection system, but shared some reservations concerning our intended method for sample testing – Borrelia bacteria are very rarely present in the blood at a high enough concentration for detection through traditional techniques. As we discussed this issue and how we could overcome this, we mentioned our plans for alternative applications. Dr Brooks emphasised how useful our sensor would be for detection of leptospirosis – a bacteria commonly detected in the blood samples RIPL test, with the paper-based aspect making it very well-suited to the developing countries where this disease prevails. We arranged a future consultation with Dr Brooks to discuss our technology further, and after conferring with the rest of the team, made the executive decision to alter the primary target of our sensor.

Collaborating with members of Public Health England combined with the analysis of collected data led us to conclude that when detecting for Lyme disease, an in vivo form of our sensor would be most appropriate. The advantages of using paper-based sensing over in vivo are the ease of use, stability and cost, however, if our system is used in clinics in developed areas, these benefits become secondary to concerns over sensitivity. An in vivo detection kit would be more easily tuneable for sensitivity than an in vitro paper-based kit, on the basis that it is more easily analysed under imaging and assay techniques, with the loss of ease of use and affordability. In developed areas, and when used by appropriately trained professionals, the benefits regarding sensitivity when using in vivo detection outweigh any negative aspects.