4-octyl itaconate modulates virulence-associated phenotypes and oxidative stress resistance in avian pathogenic Escherichia coli by targeting menB and wza

Avian Pathogenic Escherichia coli (APEC) causes substantial economic losses to the poultry industry, primarily due to its diverse serotypes and extensive drug resistance. Itaconate, a metabolite of the tricarboxylic acid (TCA) cycle, demonstrates antimicrobial activity. However, the effect of itaconate on the biological characteristics of APEC remains unexplored. Because itaconate itself has poor membrane permeability, its cell-permeable derivative, 4-OI, has emerged as a promising alternative and has attracted significant attention. This study investigates the impact of 4-OI on the biological properties of APEC, including growth, biofilm formation, antioxidant stress response, and acid-base tolerance. The results revealed that the addition of 200 μM or 400 μM 4-OI to APEC94 cultures for 14 h significantly inhibited bacterial growth. Specifically, 4-OI at concentrations of 100 μM to 400 μM markedly suppressed biofilm formation in APEC. Conversely, concentrations between 50 μM and 400 μM significantly enhanced APEC94 survival under H₂O₂-induced oxidative stress. Transcriptomic analysis demonstrated that the exogenous application of 200 μM 4-OI significantly altered the transcription of the wza and menB genes. Further investigation demonstrated that 4-OI targets the wza and menB genes, potentiating its inhibitory effect on biofilm formation and mitigating H₂O₂-induced suppression in APEC94. Furthermore, treatment with 200 μM 4-OI reduced bacterial tolerance against acid-base stress and osmotic pressure, an effect that was contingent upon the presence of the wza and menB genes. In conclusion, this study reveals the multifaceted effects of 4-OI on APEC, including its inhibitory effects on growth, biofilm formation, oxidative stress response, and tolerance to acid-base and osmotic stress. Furthermore, it demonstrates that the wza and menB genes mediate the enhancement of oxidative stress resistance in APEC induced by 4-OI.