Cellular Expression of Two Key Water Borne Pathogens towards Antimicrobial Quaternized Chitosan

Abstract

The antibacterial activity of chitosan and its quarternized derivative i.e., TMC against Salmonella typhimurium and Shigella flexneri was evaluated and an attempt to understand the mechanism of action was attempted in this study. The two key gram-negative water borne pathogens was evaluated for viability at different incubation times. Further, the integrity of the cell membranes of both bacterial cells were investigated by determining the release from cells of materials that absorb at 260 nm, electric conductivity measurements and leakage release of cytoplasmic β-galactosidase activity. Results revealed that after treatment with TMC, the electric conductivity of bacterial suspensions increased, followed by increasing of the units of average release for alkaline phosphatase (ALP) and glucose-6-phosphate dehydrogenase (G6PDH). SDS PAGE and occurrence of protein in cell free supernatant indicated that the soluble proteins decreased or disappeared in the treated Salmonella cells, demonstrating that TMC performed its antibacterial function via increasing the permeability of cell membranes. The results obtained with two water borne pathogens demonstrate the complexity of the mode of action of Quaternized biopolymer. An explanation of the antibacterial mechanism is proposed involving the cell wall disruption due to interaction between positive amino groups present in the TMC with the negatively charged bacterial cell surface. Several lines of evidence suggest that its site of action is the outer membrane, but a probability of more than one target site would explain quaternized chitosan’s antimicrobial action.