Nutrient availability and sodium pyruvate regulate the resuscitation of viable but nonculturable Staphylococcus...

Staphylococcus aureus (S. aureus) can enter a viable but nonculturable (VBNC) state when induced by citric acid at 4 °C, posing hidden risks to food safety upon resuscitation. However, the effects of nutritional conditions and sodium pyruvate (SP) on this process remain unclear. This study demonstrates that 0.5 × TSB, TSB, 2 × TSB, and SP resuscitate VBNC cells following sigmoidal kinetics, with an efficacy ranking of SP > 2 × TSB > TSB > 0.5 × TSB. SP significantly accelerated resuscitation through enhanced early enzymatic activity, a transient increase in intracellular ROS levels, and rapid ATP accumulation, accompanied by dynamic nucleic acid fluctuations peaking at 10-12 h. Furthermore, proteomic profiling and targeted inhibition analyses revealed that DNA repair and cell wall biosynthesis are critically involved in initiating resuscitation, whereas signal transduction and respiratory metabolism primarily govern post-resuscitation growth. Practically, a 5-h or 10-h pre-resuscitation in 2 × TSB successfully detected VBNC S. aureus in high pressure processing-treated apple juice during 4 °C storage, which was undetectable via direct enumeration. These findings offer mechanistic insights into pathogen resuscitation and provide practical strategies for food safety risk assessment and detection.