Induction of a viable but non-culturable state in Salmonella Typhimurium is correlated with free radicals generated by thermosonication

Abstract

The viable but non-culturable (VBNC) state in bacteria is gaining more and more attention around the world, mainly due to it is not only an important strategy for adaption to a stressful environment but also may possibly pose a threat to food safety and public health. It was found that a small subpopulation of Salmonella Typhimurium in pure culture was induced into a VBNC state during thermosonication (TS) processing in our previous study, though few known about the situation in real food and how bacteria were induced into that special state. Base on the speculation that free radicals generated during TS affected induction of VBNC, the relationship between them was investigated preliminarily. It was observed that higher intensity of TS treatment, such as higher power, elevated temperature and prolonged duration resulted in more viable S. Typhimurium cells in carrot juice been inducted into VBNC state. The observed results showed that VBNC incidence indexes were 1000 as TS treatments at 57-62 °C and 380 W for 6 min, and 53 °C and 380 W for 8-10 min were applied, indicating a 100% VBNC state of viable cells in those cases. The ESR spectra revealed three kinds of free radicals, including carbon centered (ethanol) radicals, hydroxyl radical and hydrogen protons were generated in carrot juice during TS processing. The intensity of free radicals was tied to the TS processing parameters and also influenced the occurrence of VBNC. A nonlinear sigmoidal curve of the intensity of free radicals VS the VBNC incidence index in three stages, including a slow phase (with free radicals intensity of 0-0.10), a rapid growth phase (with free radicals intensity ranged 0.10-0.14) and a final equilibrium phase (with free radicals intensity greater than 0.14), was observed and well fitted with the Boltzmann model. Moreover, the significance of free radicals generated during TS processing for induction of VBNC state was verified and confirmed with 0-200 mM sodium pyruvate. The obtained results may contribute to understand the complicated phenomenon and guide the application of TS as a decontamination technique.

Keywords: Carrot juice; Free radical; Induction factor; Salmonella Typhimurium; Viable but non-culturable.