Extracellular pH decline introduced by high pressure carbon dioxide is a main factor inducing bacteria to enter viable but non-culturable state

High pressure carbon dioxide (HPCD) has been used in food processing as a non-thermal pasteurization technology. However, the potential of HPCD to induce viable but non-culturable (VBNC) cells limits its application. The objective of this study was to explore the roles of extracellular pH of 3 (pH(ex3)) and high pressure (HP) of HPCD during VBNC induction and the underlying molecular mechanism. By using the model organism Escherichia coli O157:H7, we found that the combined effects of pH(ex3) and HP could mimic the effect of HPCD for VBNC induction. Further investigation of the individual effect of pH(ex3) and HP on VBNC induction revealed that pH(ex3) could induce a higher proportion of VBNC cells with a slower induction rate compared with HPCD, whereas HP was unable to induce VBNC formation. Notably, the cells treated by pH(ex3) and HPCD had similar morphological changes, and VBNC cells induced by pH(ex3) and HPCD had similar stress resistance characteristics. These results strongly indicated that pH(ex3) introduced by HPCD was a main factor for VBNC induction. Additionally, we found that HP played the role in accelerating VBNC formation in the process of HPCD treatment. Transcriptomic analysis revealed 85, 263 and 529 differentially expressed genes (DEGs) for HP-, pH(ex3)- and HPCD-treated cells compared with untreated ones. 59 DEGs shared by pH(ex3) and HPCD treatment might be responsible for VBNC induction, and they were mainly involved in cellular transport and localization.