Resuscitation of Pulsed Electric Field-Treated Staphylococcus aureus and Pseudomonas putida in a Rich Nutrient Medium

Pulsed electric fields (PEFs) technology was reported to be useful as a disinfection method in the liquid food industry. This technology may lead to membrane permeabilization and bacterial death. However, resuscitation of viable but non-culturable cells and sublethally injured microorganisms in food was reported to be associated with foodborne outbreaks. The main aim of this study was to investigate the possible recovery of injured PEF-treated bacteria. The PEF treatment of Staphylococcus aureus and Pseudomonas putida led to a reduction of 3.2 log₁₀ and 4.8 log₁₀, respectively. After 5 h, no colony forming units (CFUs) were observed when the bacteria were suspended in phosphate buffer saline (PBS); and for 24 h, no recovery was observed. The PEF-treated S. aureus in brain-heart infusion (BHI) medium were maintained at 1.84 × 10⁴ CFU mL^-1 for about 1.5 h. While P. putida decreased to zero CFU mL^-1 by the 4th hour. However, after that, both bacteria recovered and began to multiply. Flow cytometry analysis showed that PEF treatment led to significant membrane permeabilization. Mass spectrometry analysis of PEF-treated P. putida which were suspended in BHI revealed over-expression of 22 proteins, where 55% were related to stress conditions. Understanding the recovery conditions of PEF-treated bacteria is particularly important in food industry pasteurization. To our knowledge, this is the first comprehensive study describing the recovery of injured PEF-treated S. aureus and P. putida bacteria.