TRIAL FOR RESUSCITATION OF VIABLE BUT NON-CULTURABLE (VBNC) L. MONOCYTOGENES DUE TO THE EFFECT OF CHLORINE AND MAGNESIUM CHLORIDE (MGCL2) ON FOOD CONTACT SURFACE

Khalid Tolba*, Basma A. Hendy and Noha M. El-Shinawy

ABSTRACT

Many bacterial species including L. monocytogens have been found to exist in a viable but non-culturable (VBNC) state as a result of different stresses. VBNC cells are characterized by a loss of culturability on routine isolation by using the reference methods, which weakened their detection by conventional plate count techniques. This leads to an underestimation of total viable cells in environmental or food samples, and thus poses a high-risk level to public health. In this paper, recent findings on the VBNC state of human bacterial pathogens particularly L. monocytogens were approached. The viable cells after 30 min. of chlorine application (50, 100 and 200 ppm) were decreased by 100% [from 7 log10cfu/ml to <1 cfu/ml (0.0%)] while after 48 of resuscitation using modified detection method, the VBNC cells in case of used 50 ppm chlorine were recorded 5 log10cfu/ml (71.43%) and 2 log10cfu/ml dead cells (28.57%), however by using 100 ppm chlorine, VBNC cells were recorded 2.9 log10cfu/ml (41.43%) and 4.1 log10cfu/ml dead cells (58.57%) while by using 200 ppm chlorine, VBNC cells were recorded 2.8 log10cfu/ml (40.00%) while dead cells reached 4.2 log10cfu/ml (60.00%). Furthermore, by using MgCl2 (0.5 and 1.0 Mol), viable cells were recorded 95.71 and 85.71 and VBNC cells (4.29 and 14.29), respectively while dead cells recorded (0.0%) in both MgCl2 concentrations. Treatment with chlorine was more potent in induction of VBNC state and dead cells rather than MgCl2. PCR was able to detect inlA, prfA and hlyA virulence gens and the organism had kept the virulence genes active during VBNC state, which was revealed its existence after resuscitation. The results of the present study indicated that in all cases of treatments, whether by using chlorine or MgCl2 does not have the ability to kill Listeria monocytogens, in addition, PCR is considered a rapid method in detecting the organism and its virulence genes, while negatively samples by using PCR should be subjected to modified traditional methods of analysis. The potential influences of VBNC L. monocytogens on human health were discussed.

Keywords: VBNC, stress, resuscitation, virulence gens, PCR, human pathogens, L. monocytogens, chlorine, MgCl2.