Portable automated test for fast detection and surveillance of influenza
The PORTFASTFLU's objective is to develop and validate a rapid diagnostic test for influenza that will be used as point-of-care systems in developed and developing countries. Experts and international bodies agree that the rapid detection of influenza is vital in combating this major threat to human health. Nucleic acid analysis is the most appropriate assay scheme for both early detection and late surveillance of influenza, and is routinely used by WHO and OIE virology reference laboratories. PORTFASTFLU's consortium aims at developing and validating a Point of Care (POC) Funding scheme which will integrate sample preparation, nucleic acid amplification, microarray hybridization and fluorescent readout in a single system. Reaching this challenging objective is possible through the combined expertise of the partners and comprises access to several selected enabling technologies: microfluidics, on-chip sample preparation, advanced microarray techniques and reading systems with ultimate sensitivity and selectivity, access to reference and field virus samples. This Bio-Micro knowledge converges in one single instrument, allowing rapid detection (less than an hour) and simultaneous identification of influenza A and B viruses (typing and sub-typing) at widespread POC with only basic skills. Furthermore, the project also aims at realizing this goal in a format that requires little supervision and is integrated in a portable compact monolithic system for deployment in the field.
The consortium of 8 partners from 5 member states brings together the various expertises needed to reach these objectives.[+] Read More
The project's objective is to develop and validate a rapid diagnostic test for human influenza that will be used for surveillance and early detection of influenza and as a point-of-care tool in developed and developing countries. The diagnostic test will enable the rapid detection of influenza infection in a fast and specific way (typing and sybtyping) using a monolithic disposable cartridge placed in a compact, portable analytical instrument.
Influenza viruses which are adapted to human and have caused pandemic or epidemic waves so far are characterised as H1, H2 or H3 sub-types in combination with N1 or N2 sub-types. These viruses are the major targets for the detection tool to be developed under this application. Indeed, they cause several thousands of deaths each year in Europe and there is a real need for a test providing fast identification of the causative agent and thus contributing to the improvement of treatment and surveillance.
However, avian influenza virus can also be transmitted to humans and may cause severe disease. Therefore, the PORTFASTFLU consortium has decided to also include the H5, H7 and H9 avian sub-types in the new tool to be developed. For the currently circulating strain H5N1 lineages of virus, it is clear that a miniature biosensor allowing rapid detection of the virus would help contain the infection and combat the threat of disease at a very early stage. For persons having flu-like symptoms that are admitted to hospitals in regions with H5N1 cases epidemics in poultry or wild birds, it is vitally important to make a rapid point-of-care diagnosis in order to implement recommended control measures as fast as possible.
The technology that PORTFASTFLU intends to assemble in a single machine would provide essentially the same information as at WHO laboratories within 30 minutes to 1 hour after the start of sample analysis (e.g. throat or oro-nasal swabs). Furthermore, the system is designed such that it is easy to use, does not require highly qualified personnel, is robust and has an inbuilt GPS system and an on line data transmission system. In the design of the new tool, the critical points will be the selection of the appropriate sequences for effective priming and probing and also the miniaturisation of the RNA preparation.
For the identification of appropriate sequences, a comprehensive bioinformatics approach will be carried out and selected sequences will be validated with the nucleic acid tests that are already available.
The consortium is aiming to develop a single Influenza diagnostic technology using two different sample preparations routes embodied as cartridges compatible with the same readers. Within the project, two Point Of Care systems will therefore be developed as proof of concept, each one using a different cartridge but sharing common hardware and software.
The first one will use a cartridge for PCR detection of bacteria which will be embedded in a compact POC instrument with the highest capacity for functionality including multiplex amplification and real time hybridization/melting monitoring.
The second one, relying on a a solid RNA capture matrix embedded in a disposable fluidic chip, will be easier to handle in the field but more risky technologically (as relying on an unproven concept),will be built as a field battery-powered system with functionalities to be defined along the project.
The final validation will emphasize on the system that fits the best the requirements of users in terms of sensitivity, reliability, cost, portability, and ease of use.
We believe that this novel diagnostic tool will speed up diagnosis and subtype specification of influenza virus considerably: the test is rapid and aimed for on site as well as laboratory use by personnel with only basic skills. The test will allow a far more detailed and real-time monitoring of influenza and its aetiology, which still is not well understood yet. Undoubtedly this will also provide faster and more accurate information to influenza vaccine manufactures and allows a more sound prediction of the required vaccine strains.
Figure 3. Schematic representation of the PORTFASTFLU Point-if-care system for rapid diagnostic test for influenza
The processes of miniaturisation proposed here plus its paralleling, micro-integration and mass production have the potential of implementing major cost reductions that will broaden the use of these types of devices and the combined portable reader, making these natural parts of the diagnostic system for potential use in health care, but also in the food chain control. Their main impact can be identified as cost reduction, reduced analysis time and increased throughput.
So far, different technologies have been used for biosensor fabrication. So far, few have gone to full integration of the biological contents and measurement system.
The successful development of a full diagnostic kit, component and system as described in this project will impact the low cost diagnostic test market place and create a standard for future similar tests, imposing a major reference in the field. Several items are particularly important technologically, even if considered separately from the others: the diagnostic influenza kit, not yet demonstrated; the sample preparation, which can be applied in a number of other major needs, the compact, fast and non-supervised hybridization and readout system with communications and GPS capacities.
The PORTFASTFLU proposed technology will bring about three main improvements:
The ability to process a sample, analyse, and relay results to appropriate authorised reference labs using one easy-to-use portable system would be of enormous benefit to Influenza surveillance. The ability to immediately identify the causative agent of avian disease on commercial premises could have enormous impact on the financial economic burden associated with outbreaks of avian influenza, and result in a dramatic decrease in the time required to implement the necessary control measures. The benefits of this would be particularly visible in developing nations currently tackling avian influenza, such as Central African countries and nations in South East Asia. Similarly, the ability to instantly detect and subtype viruses in human disease cases would have enormous benefit, both in the fight against recurrent seasonal influenza, and for use in the event of a major epidemic or pandemic of influenza.