Sensitive peptIde- and apta-assays for on-site dEtectioN of bActerial cells and spores in milk and meat samples – Siena

Food poisoning is the first cause of hospitalization over the world. Current diagnostic techniques for foodborne bacterial pathogens are either expensive and time-consuming, or need isolated genetic materials and sophisticated instruments. They cannot be applied on-site to allow food traceability. These considerations apply to Campylobacter, which is the most common bacterial cause of human gastroenteritis in the world, and Bacillus cereus, the primary microbe contaminating baby food and the third agent responsible for collective foodborne outbreaks in Europe.
We propose an innovative and ambitious project to provide a low-cost assay based on the recognition properties of small peptides and aptamers, and an electrochemical sensor connected to a portable instrument for on-site food analysis. Electrochemical biosensors harness the specificity and sensitivity of biological systems in small, low cost devices providing a powerful alternative to conventional methods. Moreover, electrochemical biosensors enable analysis and screening of foodstuffs during all phases of production providing efficient tools to ensure food safety.
The first strong point of this project is the design and selection of new recognition elements for targeted pathogens. Large-scale affinity screening based on phage display will be employed to select specific peptides that bind surface epitopes of targeted bacteria. A colorimetric test based on gold nanoparticles will also be used to optimize previously selected specific aptamers. The obtained set of preselected peptides and aptamers with good affinity to our targets will be grafted on a biochip and their performance for bacterial detection will be evaluated simultaneously using a high-throughput optical platform surface plasmon resonance imaging (SPRi). Finally, selected peptides and/or aptamers with the highest affinity will be used to construct a portable and multiplex electrochemical biosensor for diagnosing foodborne pathogens. The second strong point of the project concerns the original production of the electrochemical sensing platform. The device will be entirely manufactured using the fused deposition modelling 3D-printing technology, which offers the best price/performance ratio for prototyping of small-scale production. The third strong point of the project will be the validation of the electrochemical device. Validation tests for Campylobacter detection in poultry meat will be performed on C. jejuni, C. coli, C. lari, C. upsaliensis and C. fetus, as these strains caused more than 90 % of all human campylobacteriosis. Tests for Bacillus cereus detection in milk will be performed using the INRAE laboratory collection, which contains strains isolated from human milk banks and foodborne outbreaks (>500 strains). The assay will be optimized for control of infantile formula and human milk banks for the presence of B. cereus cells and spores. Currently there is no such portable test for pathogen diagnostics in food samples in EU. Our electrochemical biosensor will enable in-situ identification of pathogens causing foodborne illness, which will improve food safety.