A high level of cell-contact cytotoxic activity was consistent with the ability to induce HEp-2 HTS cell death. The Pearson linear correlation test revealed positive correlations between the Apoptotic Index of infected HEp-2 cells at 24h, cell-contact cytotoxicity (r = 0.59, P < 0.01), and cell-contact hemolytic activity (r = 0.77, P < 0.01) and the Invasion Index (r = 0.63, P < 0.01). The results indicated that S. marcescens strains can produce cell-bound cytotoxins that increase ability to invade epithelial cells and induced their apoptosis. A number of microbial factors that can trigger the induction of apoptosis of host cells have been identified. Previously, Carbonell et al.

[4] observed that a cytotoxic toxin isolated from clinical strain was bound to the CHO cell surface, without being internalized and next was able to trigger changes in intracellular metabolism of the cells and to induce cell death by apoptosis. Hertle [3] suggested that ShlA induced irreversible vacuolation with subsequent lysis of epithelial cell lines and erythrocytes. Massive disruption of the host cell membrane by pore-forming toxins disturbed cellular homeostasis and resulted in cell death by apoptosis [5]. In the present study we observed that 9 (30%) strains induced high Apoptotic Index of epithelial cells above 60% and low cell-contact activity. High apoptotic activity could be associated with extracellular toxins produced by the strains. We have previously observed that bacterial culture supernatant 9 of 20 S. marcescens strains used in this study revealed cytotoxic activity to HEp-2 cells at 24h incubation [8].

Previous studies suggested that S. marcescens strains produce RNase that display potent cytotoxic activity [17]. Shimuta et al. [9] reported that S. marcescens strains produced extracellular phospholipase A (PhlA) that revealed hemolytic and cytotoxic activity. PhlA induced destabilization of target cell membranes by directly hydrolyzing their phsphlipids. The pore induced by the toxins results in increasing cytosolic Ca+2 which could be the signal for the initiation of apoptosis. Carbonell et al. [4] isolated extracellular cytotoxic enterotoxin from a clinical isolate of S. marcescens which was highly cytotoxic to CHO cells but did not reveal hemolytic activity, which suggested that the cytotoxin is distinct from S. marcescens hemolysins.Phagocytes, either resident in tissues or circulating in blood, contribute to the primary line of innate defence against bacterial pathogens by providing their removal and destruction at the level of epithelial barrier [5]. Some bacterial pathogens induce the death of immune cells to subvert normal host defence mechanisms, for invasion to deeper layers of the Cilengitide tissue.