The biochemical characteristics of viable but nonculturable state Yersinia enterocolitica induced by lactic acid stress and its presence in food systems.

The viable but nonculturable (VBNC) state is adopted by many foodborne pathogenic bacteria to survive in adverse conditions. This study found that lactic acid, a widely used food preservative, can induce Yersinia enterocolitica to enter a VBNC state. Y. enterocolitica treated with 2 mg/mL lactic acid completely lost culturability within 20 min, and 10.137 ± 1.693 % of the cells entered a VBNC state. VBNC state cells could be recovered (resuscitated) in tryptic soy broth (TSB), 5 % (v/v) Tween80-TSB, and 2 mg/mL sodium pyruvate-TSB. In the VBNC state of Y. enterocolitica induced by lactic acid, the intracellular adenosine triphosphate (ATP) concentration and various enzyme activities were decreased, and the reactive oxygen species (ROS) level was elevated, compared with uninduced cells. The VBNC state cells were significantly more resistant to heat and simulated gastric fluid than uninduced cells, but their ability to survive in a high-osmotic-pressure environment was significantly less than that of uninduced cells. The VBNC state cells induced by lactic acid changed from long rod-like to short rod-like, with small vacuoles at the cell edges; the genetic material was loosened and the density of cytoplasm was increased. The VBNC state cells had decreased ability to adhere to and invade Caco-2 (human colorectal adenocarcinoma) cells. The transcription levels of genes related to adhesion, invasion, motility, and resistance to adverse environmental stress were downregulated in VBNC state cells relative to uninduced cells. In meat-based broth, all nine tested strains of Y. enterocolitica entered the VBNC state after lactic acid treatment; among these strains, only VBNC state cells of Y. enterocolitica CMCC 52207 and Isolate 36 could not be recovered. Therefore, this study is a wake-up call for food safety problems caused by VBNC state pathogens induced by lactic acid.