Final Report Summary - BUGCHECK (A rapid hand-held analyser for control of microorganisms in the complete meat supply chain) Foodborne pathogens in processed ready-to-eat products pose a serious threat to consumers with compromised immune system. Sensitive, specific and rapid detection of such pathogens is thus essential at production level to prevent their entrance into the human food chain. Conventional microbiological detection methods simply take too long (two to seven days) to detect and identify pathogens in food and no real time data is available. Traditional testing methods, such as enzyme-linked immunosorbent assay (ELISA), are relatively costly and time-consuming. Traditional methods require the taking of a product sample, its posterior culturing until sufficient microorganisms have been generated to enable ready detection on culture plates. While a number of methods such as polymerase chain reaction (PCR) may provide faster detection (6-12 hours) they involve complex procedures and highly specialised trained personnel. In today's modern food supply chain, products enter and leave the market within two to three days. Slow traditional analytical methods are clearly deficient as they enable contaminated meat products to reach the market, resulting in human disease and even mortality. Moreover, most analyses need to be carried out in large analytical laboratories as the required instrumentation is expensive and requires highly qualified staff. Only very large farms and slaughterhouses can afford to perform regular, on-site, microbiological checks. The project investigated, developed and validated a multi-analyte platform based on biosensor technology for the detection of the most common pathogens occurring in the meat industry (Campylobacter, Salmonella and Escherichia Coli), including the most common strands of each. A biosensor-based approach presents a promising and sensitive alternative tool to detecting low numbers of cell within a few minutes (with no need for enrichment steps) as opposed to days. During first steps of the project, a market survey was carried out. The objective was to ensure that the technology developed corresponds to the needs of the market place, the price sensitivity, as well as the acceptance of the proposed equipment into the market. A questionnaire and a cover letter introducing the project and its objectives were drafted and with the help of all partners were translated into different languages: English, German, Hungarian, French, Italian, Spanish and Czech. The questionnaire could be filled in on internet web page. The questionnaire was advertised on several on-line specialised magazines, a massive e-mailing was done, and a telephone survey was carried out to gain an improved and more in-depth overview. The most important conclusion gathered with the market survey was that the price of the analysis have to be around EUR 10 and the maximum result turn around 1 day. Based on this market survey as well as the experience of the partners, the general input requirements were defined. The next step in the project was to define the technical and biological requirements of the proposed system. Based on current epidemiological data available for incidence of foodborne disease in humans within the European Union, the pathogenic microorganisms to be detected was Salmonella, Campylobacter and verocytoxigenic strains of Escherichia coli. Then, suitable antibodies in the three selected microorganisms were identified taking into account the specificity to recognise the corresponding pathogens, cross-reactivity towards other pathogens as well as the capability to be used for immobilisation techniques. Once the definition of the specifications was determined and the suitable antibodies purchased, the design and development of the four main parts of the BUGCHECK prototype started: the optimisation of the immunofunctionalisation protocols, the interdigitated microelectrodes, the 'plug and play' cartridge and the electrochemical impedance spectroscopy (EIS) instrumentation. At the end of the project, it was possible to detect Escherichia coli and Salmonella in phosphate buffer solutions, using electrochemical impedance spectroscopy. The detection limit was strongly conditioned by the microelectrode geometry, reaching the best detection limits (around 104 CFU mL-1) at the 10 x 10 interdigitated structures. On the other hand, when a more complex sample matrix, such as culture medium, was used, the detection was not possible at any concentration level. The reason is very likely to be found in the large amounts of organic matter that can readily adsorb on the chip surface, thus blinding the sensors. One possible way to overcome these matrix effects may be the use of paramagnetic particles to extract the target pathogen from the sample and then transfer them to a cleaner environment, such as a phosphate buffer solution. Related with project results exploitation, the partners of the consortium agreed that the overall BUGCHECK system had very low commercial value and has no sense to commercialise it, however, there are some final results potentially exploitable: A) immobilisation procedure B) biosensor software package C) plug and play cartridge system D) microelectrodes on silicon and glass E) validated method of the measurement. All SME partners were formally asked to express their interest in BUGCHECK project results and only three of them (BVT, BST and Aromics) were interested in one or more of these exploitable results. An internal agreement was arranged between these SME partners. Related with dissemination of the results, a website has been set up containing the public synopsis sheet, the objectives of the project and some general information about the partners. Regarding the communication among the partners, the same website contains a private password-protected area where all the documents of the project are available. The public website is at http://bugcheck.cric-projects.com Related documents Final Report - BUGCHECK (A rapid hand-held analyser for control of microorganisms in the complete meat supply chain)
