Inhibitory activity of colicins against clonal diarrheagenic Escherichia coli strains and in vitro cytotoxicity of nisin, pediocin, colicins E1, E3, E6, E7, and K, against vero monkey kidney cells and simian virus 40-transfected human colon cells.

摘要

Twenty-four colicins were evaluated for inhibitory activity against 27 diarrheagenic E. coli strains of serotype O15:H-, O26:(H11, H-), O111:(H8, H11, H-), including serotype O157:H7, representing diarrheagenic E. coli (DEC) clones 3, 4, 8, 9 and 10 that are commonly associate with shiga-like toxin-production. Overlay techniques were used to assess inhibition on Luria agar and Luria agar supplemented with 0.25 {dollar}mu{dollar}g/ml mitomycin C to induce colicin-production. Group A colicins were inducible and had very broad inhibitory profiles, whereas group B colicins were often not inducible. Sensitivity to colicins by the DEC strains was in the following order; DEC clone 9 {dollar}>{dollar} 4 {dollar}>{dollar} 3 {dollar}>{dollar} 10 {dollar}>{dollar} 8. Strain 8C from clone 8 was resistant to most test colicins. In turn, the 27 DEC strains were evaluated for colicinogeny as above. Ten (37%) of the DEC strains tested produced colicin which partially explains why some DEC strains were insensitive to standard colicins. The comparative in vitro cytotoxicity of nisin, pediocin, colicin (Col) E1, E3, E6, E7 and K, was evaluated against Simian virus 40-transfected human colon (SV40-HC) cells and Vero monkey kidney cells grown as monolayers in Dulbecco's modified Eagle's medium supplemented with 10% (v/v) calf serum (35{dollar}spcirc{dollar}C, 10% CO{dollar}sb2,{dollar} humidified incubator). Trypan blue staining and cellular membrane potential changes using Interactive Laser Image Cytometry (ILIC) analysis with DiOC{dollar}sb5{dollar}(3), a potential-sensitive dye, were used as cytotoxic indices. Bacteriocins at final concentrations of 170, 350 and 700 AU/ml were evaluated. Nisin, pediocin, and Col E6 were the most toxic bacteriocins; the SV40-HC cells demonstrated greater sensitivity. ILIC analysis was more discriminating than trypan blue staining; it distinguished normal Vero cells as less sensitive than SV40-HC cells. There is potential for use of ILIC analysis for studying the effects of bacteriocins on mammalian cells in culture. Bacteriocins from food-grade bacteria can be toxic to mammalian cells as has been reported for colicins. Col E1, E3, E7 and K exhibited least toxicity indicating the potential for use of such colicins as biocontrol agents against pathogenic E. coli strains.