ETH Zurich, Institute of Food, Nutrition and Health Schmelzbergstrasse 7 B36

1. The enemy of our enemy is our friend; utilizing the natural killers of bacteria to improve food safety Denyes J.M., Krylova S., Marti R., Klumpp J., and Loessner M. J. ETH Zurich, Institute of Food, Nutrition and Health Schmelzbergstrasse 7 B36 8092 Zurich, Switzerland

2. Overview of presentation Beards G. 2008 Bacteriophages; natural killers The coolest video ever Attempting to decipher the mysterious phenomenon of bacterial recognition and adsorption Making tail fibers work for us, and then putting them to good use Acknowledgements D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

3. Bacteriophages; natures bacterial killer • Viruses which infect only bacteria • Most numerous organisms on the planet, estimated at 1032 • Found in absolutely every environment where bacteria are found • Either lytic or lysogenic • Kill the bacteria immediately • Insert their genome into that of the bacteria, and wait for better times to reproduce • Used in detection, typing, and bio-control of bacteria in many environments D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

4. D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

5. Eyes, ears, nose, hands, and feet… Kometani and Matsui, 2005 Raaijet al., 2001 Bartualet al., 2010 D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

6. Infection and adsorption; unique profiles! Strain 1 2 3 4 5 6 7 Propagation strain Sequencedtypestrain Biofilmstrain C. malonaticus C. Turicensis C. sakazakii C. sakazakii C. sakazakii C. sakazakii C. sakazakii Infection - +++ +++ +++ - ++ - D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

7. Leimanet al., 2010 D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

8. Making them work for us; GFP fusions pH 7 pH 4 LTFS.typhimurium DT1755 STF

9. Receptor analysis Outer surface • Deletion mutants of outer membrane proteins • Test infection, adsorption and binding with the proteins • Identified OmpC for the Salmonella phage • Sequence genes from positive and negative strains • 3 amino acid change =Positive to negative Periplasm Modified from Prasanthet al., 2012 D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

10. Putting them to good use • Biotinylation • Coupled to magnetic nanoparticles: specific • Specific and sensitive detection of contamination food after a 5 hour enrichment • 4 days shorter than other methods D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

11. In summary Bacteriophage are ubiquitous and present many advantageous traits to exploit Adsorption and infection are very different processes and phage can typically recognize a broader range of bacteria than they can infect Producing separate GFP tail fiber fusions allow for visual detection of bacteria When coupled with a solid phase immobilization instrument or nano-particle system specific, sensitive, and rapid detection of bacteria from contaminated food is possible Development of these tools is on-going D-HEST/ Institute of Food Nutrition and Health/ Food Microniology

12. Acknowledgments Azerbaijan Young Scientists, Postgraduate and Masters Union (AYSPMU) The Ministry of Youth and Sport of Azerbaijan Republic Azerbaijan Academy of Sciences Dr. Loessner, Dr Klumpp, Dr. Stephan, Dr. Miller Food Microbiology lab and all the students who have worked on the project This project was funded by Grant AiF 16756 N Förderung der industriellenGemeinschaftsforschung und -entwicklung (IGF) BundesministeriumfürWirtschaft und Technologie, Deutschland

13. D-HEST/ Institute of Food Nutrition and Health/ Food Microniology