Bacteriophage-mediated restoration of epithelial integrity, DNA damage response, and transcriptomic homeostasis in colon organoids exposed to ETBF and pks + E. coli

Colorectal cancer (CRC)-associated pathobionts such as Enterotoxigenic Bacteroides fragilis (ETBF) and colibactin-producing (pks+) Escherichia coli disrupt epithelial barrier function, promote chronic inflammation, and induce genotoxic stress. To investigate how these pathogenic interactions with the host epithelium can be modulated, we employed patient-derived normal human colon organoids as an ex vivo model. Two lytic bacteriophages, Bac-FRP-3 (targeting ETBF) and Esc-COP-23 (targeting pks + E. coli), were applied to bacterial cultures, and the resulting phage-conditioned supernatants were used to challenge organoids. Exposure to ETBF and pks + E. coli induced strong pro-inflammatory transcriptional responses, DNA damage signaling, and epithelial disruption, whereas phage treatment attenuated these effects. Transcriptomic profiling revealed restoration of inflammatory and DNA damage-related gene expression, while confocal imaging confirmed recovery of epithelial junctional integrity and reduced DNA double-strand breaks. These findings highlight phage-mediated modulation of host-pathobiont interactions and provide mechanistic insight into how targeted viral predation can reshape pathogenic impacts within the human intestinal environment.