Intracellular pH as an indicator of viability and resuscitation of Campylobacter jejuni after decontamination with lactic acid

The aim of the study was to determine intracellular pH (pHi) as an indicator of the physiological state of two Campylobacter jejuni strains (603 and 608) at the single cell level after bactericidal treatment with lactic acid (3% v/v lactic acid, pH 4.0, 0.85% w/v NaCl) and during recovery and survival using Fluorescence Ratio Imaging Microscopy (FRIM). After exposure to lactic acid solution a decline in pHi to 5.5 (FRIM detection limit) was observed in the majority of cells (75–100%) within 2 min. The enumeration data revealed that after 2 min of lactic acid exposure, approx. 90% of the initial population became unculturable. In the following 10 min of exposure, a further decrease in the cell count was observed resulting in 3.53 and 3.21 log CFU/ml reduction of culturable cells at the end of the treatment. On the contrary, the FRIM results revealed that the subpopulations with pHi > 5.5 increased between 2 and 12 min of exposure to lactic acid. Removing the acid stress and incubating the cells suspension under the more favourable conditions resulted in an immediate increase in cell population with pHi > pHex for both C. jejuni strains. Further 24 h incubation at 37 °C resulted in increased pHi and colony count (recovery study). On the contrary, 24 h incubation at suboptimal temperature of 4 °C, showed pHi decrease to pHex = 6.0 (no pH gradient) in the whole population of C. jejuni cells. Rather than dying, cells exposed for longer time (72 and 120 h) to 4 °C increased the subpopulation of the cells with positive pH gradient, mostly comprised of the cells with ΔpH > 0.5, indicating the ability of C. jejuni cells to regulate their metabolic activity under suboptimal conditions.