Cinnamon essential oil synergizes with sodium hypochlorite to suppress VBNC formation in Escherichia coli O157:H7 by modulating the temporal expression of arcA/relA-associated stress regulators.

Chlorine stress induces Escherichia coli O157:H7 (E. coli O157:H7) to enter the viable but non-culturable (VBNC) state, in which the bacteria evade traditional detection methods and exhibit enhanced tolerance, posing a serious threat to food safety. In this study, the E. coli O157:H7-WT, E. coli O157:H7-ΔarcA, and E. coli O157:H7-ΔrelA knockout strains were used. By combining checkerboard analysis, viability state detection, and determination of key gene expression throughout the whole process under sodium hypochlorite (SH) stress, cinnamon essential oil (CEO) stress, and combined SH and CEO stress, the combined antibacterial effect of the two agents was evaluated, the inhibitory effect of adding CEO to SH on VBNC state formation was clarified, and the underlying inhibitory mechanism was explored. The results showed that when SH and CEO were used in combination, the fractional inhibitory concentration index (FICI) was 0.5, indicating a synergistic effect. The addition of CEO to SH enhanced its bactericidal activity and reduced and delayed the number of E. coli O157:H7 cells entering the VBNC state as well as the process of VBNC state formation. After CEO was added to SH, the expression levels of the arcA and relA genes were reduced by 2.06-2.13-fold, and the peak gene expression was delayed by 20 min. In conclusion, the addition of CEO to SH reduced the ability of E. coli O157:H7 to enter the VBNC state and enhanced bactericidal activity by suppressing the expression of the arcA and relA genes and delaying their temporal expression. This study explored the synergistic effect and mechanism of natural active substances and chlorine-containing disinfectants from the perspective of dual-target regulation involving ArcA and the stringent response, providing a theoretical basis and perspective for improving disinfection efficiency in food processing and developing novel prevention and control strategies against VBNC bacteria.