Abstract
Among the challenges agriculture and food production has to face in the future, is the long-term management of sustainable food production for the growing world population. A crucial factor in this respect is the control of diseases and improvement of animal health as a basis for safe, efficient, and high-quality production. The impact of disease on global animal production and thus food security are measured in the £billions. Globalisation and climate change are driving the unprecedented increase of emerging and re-emerging animal diseases and zoonotics. Therefore improving animal health systems by providing tools to enable early detection and control of economically important global livestock diseases is the most effective response to this alarming situation. Foot-and-mouth disease (FMD) is a livestock disease of great economic importance, caused by the highly contagious foot-and-mouth disease virus (FMDV). The disease circulates in 77% of the global livestock population, costing ~between $5 -21 billion in production losses and control measures. The greatest overall economic loss by region is China totalling more than 2 billion USD, whereas the direct financial impacts as a result of production losses are greatest in Africa with an estimated loss of $815 million. Countries where FMD is endemic pose a great threat to disease free countries such as the UK for example the 2001 outbreak cost more than £8bn to control. FMD results in these huge economic losses through production losses (producers, markets, slaughterhouses, food processors, and consumers), disease eradication losses (quarantine enforcement, euthanisation, disposal of carcases, compensation, disinfection) and trade losses (restriction of trade to FMD free countries). Routine laboratory tests are widely used for FMD diagnosis and although relatively quick, the speed of result reporting is confounded by the time delays associated with sending samples from the farm to the laboratory. These delays have direct impacts upon the quality of evidence that can be used to support immediate diagnostic decisions in the event of suspect cases of FMD. The lack of a quick test result is particularly important for suspect FMD cases in sheep and goats, since diagnosis based solely on clinical signs can be very challenging in these species. These factors have driven the development of rapid tests which can be used in situ. In fact, the use of such rapid diagnostic assays was recommended in major reports following the UK 2001 FMD outbreak. Simple pen-side diagnostics which can be deployed on farm to rapidly detect viral antigen have been developed in the form of lateral flow devices (LFD's) for detection of FMDV, however the sensitivity of these tests is less than that of molecular methods. Therefore, research was focused on transfering real-time rRT-PCR into the field, but with the cost of most platforms being expensive and the need for decontamination of the equipment, emphasis has shifted to alternative assays such as loop-mediated isothermal amplification (LAMP). However, at present LAMP still requires expensive reagents and equipment, multiple pipetting steps and a skilled operator to perform and interpret the test. Safe-guarding of animal health through prevention and control of animal diseases is therefore of paramount importance for sustainable food production and veterinary public health. This project will see the development and validation of the first 'diagnostic FMDV sensing spray' which we anticipate can be applied non-invasively to the mouth, feet, nasal and teat area of animals suspected of displaying clinical signs of FMDV. Using an innovative approach a non-toxic dye which can be detected by the naked eye will be released from nanoparticles which only respond when they either come into contact with the whole FMDV virus or with enzymes released by the virus.
Technical Summary
Previous research at The Pirbright Institute has shown that, in principle, proteases (3Cpro and Lpro) produced by foot-and-mouth disease virus (FMDV) could be exploited as diagnostic tools to confirm the presence of FMDV. It is our hypothesis that we can transfer this knowhow and other theoretical approaches to the creation of novel mesoporous silica nanoparticle (MSN) sensing systems which exclusively respond to the presence of FMDV proteases by releasing a visible dye. Guided by our existing knowledge and expertise within this area, we will first determine the correlation between the titre of FMDV and the 3C and L protease activity using cattle epithelial samples from a range of serotypes. Specificity of this protease action will be determined using related viruses (objective 1). Next (objective 2) we will focus on the creation and laboratory validation of three MSN sensing platforms, the first and second of which will involve capping of the MSN pores (to prevent dye release) with peptide sequences representing the recognition sites for 3Cpro and Lpro respectively, whilst the third will involve capping the MSN pores with Mab D9 (recombinant O serotype monoclonal antibody) linked to the MSN via a modified peptide sequence representing the VP1 G-H loop. In the presence of either 3Cpro or Lpro the peptide sequences capping the pores on the first and second MSN platforms will be cleaved and the dye released. In the case of the third platform, upon encountering FMDV the MAb due to its higher affinity to the whole virus than the modified peptide will disassociate from the pore releasing the dye. Finally we will take the most promising MSN forward for formulation into a non-toxic, non-invasive diagnostic spray and validate this for diagnostic specificity and sensitivity using archival field and experimental clinical samples (Objective 3).
Planned Impact
Among the challenges agriculture and food production has to face in the future, is the long-term management of sustainable food production for the growing world population. A crucial factor in this respect is the control of diseases and improvement of animal health as a basis for safe, efficient, and high-quality production. The impact of disease on global animal production and food security are measured in billions of pounds. Foot-and-mouth disease (FMD) is a livestock disease of great economic importance, caused by the highly contagious foot-and-mouth disease virus (FMDV). The disease circulates in 77% of the global livestock population, costing ~between $5 -21 billion in production losses and control measures. The greatest overall economic loss by region is China totalling more than 2 billion USD, whereas the direct financial impacts as a result of production losses are greatest in Africa with an estimated loss of $815 million. Countries where FMD is endemic pose a great threat to disease free countries such as the UK for example the 2001 outbreak cost more than £8bn to control. FMD results in these huge economic losses through production losses (producers, markets, slaughterhouses, food processors, and consumers), disease eradication losses (quarantine enforcement, euthanisation, disposal of carcases, compensation, disinfection) and trade losses (restriction of trade to FMD free countries). Safe-guarding of animal health through prevention and control of animal diseases is therefore of paramount importance for sustainable food production and veterinary public health. An integral part of any prevention and control strategy are powerful diagnostic tools that can be implemented in a wide range of settings and in particular at the point of care application. Diagnostic tests developed within this project will present a new level of simple, non-invasive veterinary diagnostic tool for FMD which impairs food production in many countries. In consequence, the common market will be strengthened and the losses caused by animal disease outbreaks reduced. Immediate beneficiaries will be national, regional and international laboratory scientists, stakeholders, decision makers and other key beneficiaries with an interest in animal health and food security (e.g. The Food and Agriculture Organization of the United Nations, The European Commission for the Control of Foot-and-Mouth Disease (EuFMD), the OIE (World Organisation for Animal Health, and the Directorate-General for Health and Consumers [DG SANTE]). The Pirbright Institute (TPI) is fully committed to engage the public's attitudes to science and the impact of the project will also be publicised by communication with the farming community and the public through our web page, TV and radio, workshops, training courses and exhibitions. Our results will also be of great interest to the academic community studying mechanisms to permit rapid, non-invasive disease detection and, where possible, will be made available through peer review publications and scientific meetings. TPI fully embraces the need for its science to underpin the needs of a range of activities undertaken by commercial/industrial Science Companies. The commercial exploitation and intellectual-property potential of the research will be maximised through regular interactions between the Business Development Managers at TPI. If our technologies are found to be commercially viable, a strategy for intellectual-property rights will be implemented. TPI undertakes high quality research and is committed to the development of highly skilled scientists. The research investigators will receive excellent training in the fields of virology and chemistry, thereby contributing to the wider training, innovation, skills and capability of the UK science base with consequent boosting to science-based industries and the UK economy.