Trans-cinnamaldehyde triggers stringent response-mediated virulence attenuation in pathogenic Escherichia coli

Antimicrobial resistance in Escherichia coli remains a critical threat in both human and veterinary medicine, demanding innovative strategies beyond traditional antibiotics. Here, we reveal that trans‑cinnamaldehyde (t‑CA), a natural phenylpropanoid, not only inhibits bacterial growth but also actively reshapes transcriptional programs through the RelA‑mediated stringent response. Using RNA‑seq, we demonstrate that sub‑inhibitory t‑CA (0.25 ×MIC) triggers broad transcriptional remodelling, suppressing key virulence pathways such as iron acquisition, quorum sensing, and acid resistance - effects that were not pronounced in a relA-deficient background. Consistently, t-CA induced RelA-dependent accumulation of the stringent response alarmone, (p)ppGpp, linking stress signalling with repression of virulence determinants. Functionally, t-CA impaired extracellular matrix production and significantly reduced biofilm biomass and metabolic activity. In host-relevant assays, t-CA showed moderate cytotoxicity in mammalian cell lines and attenuated LPS-driven inflammatory cytokine output in Peripheral Blood Mononuclear Cells (PBMCs) model. Finally, post-infection administration of t-CA improved survival in the Galleria mellonella infection model. Together, these results identify stringent response signalling as a key axis underlying t-CA-driven virulence attenuation and support t-CA as a candidate antivirulence adjuvant for combating difficult-to-treat E. coli infections.