Fluorescence and Electron Microscopy of Fusobacterium nucleatum-Induced Autophagy.

Fusobacterium nucleatum, an opportunistic oral anaerobe, has emerged as a key microbial contributor of colorectal cancer (CRC) progression and chemoresistance through its modulation of host cellular pathways. Autophagy-a conserved degradation process-has been identified as a central mechanism by which F. nucleatum enhances tumor cell survival. Here, we describe a standardized platform for detecting and quantifying F. nucleatum-induced autophagy in CRC cells using immunofluorescence microscopy with an mRFP-GFP-LC3 reporter to measure autophagic flux, complemented by ultrastructural analysis of autophagosomes via thin-section transmission electron microscopy. This protocol provides a robust and reproducible framework for investigating host-microbe interactions in CRC and can be readily adapted to evaluate autophagy induction by other bacterial species.