ISPHAGE · Deciphering the molecular architecture and infection mechanisms of corynephages within native host environments

Antibiotic resistance is a major global health threat, creating an urgent need for alternative therapeutics. Phages, viruses that infect and kill bacteria, are promising candidates, but their biomedical use requires detailed knowledge of infection and replication mechanisms at atomic resolution. Corynephages infect bacteria of the Corynebacteriales order, including important pathogens such as Corynebacterium diphtheriae and Mycobacterium tuberculosis. Despite their relevance, these phages remain entirely unexplored at the structural and molecular level. Their fragile and flexible structure has so far prevented structural studies using cryo-electron microscopy (cryo-EM).The ISPHAGE project aims to overcome these limitations and determine, for the first time, the complete atomic structure of a corynephage. This will deliver unprecedented insights into their infection strategies and replication cycle in the natural host. To achieve this, I will establish an integrated cryo-EM pipeline combining single particle analysis (SPA), cryo-electron tomography (cryo-ET), subtomogram averaging (STA), and cryo-focused ion beam (cryo-FIB) milling. Furthermore, I will implement innovative computational approaches to address protein flexibility during STA, currently a major barrier to achieving high resolution in cryo-ET.By delivering the first de novo atomic details of corynephages and visualizing infection events in situ, ISPHAGE will significantly advance our understanding of phage biology. The project will strengthen my career as an independent investigator through advanced training and knowledge transfer, while paving the way for innovative antibacterial strategies. In this way, ISPHAGE will contribute both to fundamental science and to the global fight against antimicrobial resistance, a critical priority for Europe and worldwide health.