Laser interferometric and cultivation methods for measurement of colistin/ampicilin and saponin interactions with smooth and rough of Proteus mirabilis lipopolysaccharides and cells

Abstract

Laser interferometry is commonly used in permeability studies of soluble substances. In this study a modification that allowed testing partially insoluble mixtures is presented. The modification relies on the measurement of diffusion from 1% agarose gel. As a model for this study, two Proteus mirabilis strains were used that differ in polysaccharide content: smooth P. mirabilis S1959 strain and its Re-type mutant, strain R45. By laser interferometry and precipitation it is shown that R45 lipopolysaccharide is more effective in binding colistin. It has been shown with the laser interferometric method that saponins, which are detergent-like substances of plant origin, partially enhance the interaction of colistin with the S and Re types of P. mirabilis. These results were confirmed with whole cell Proteus studies. The saponin partially inhibited the growth of the S and Re P. mirabilis strains at doses of 31–500 μg/ml. A sub-inhibitory dose — 15 μg/ml of saponins alone do not reduced the numbers of P. mirabilis S1959 and R45 cells. However, the presence of colistin or amipicillin and 15 μg/ml of saponins reduced the amount of P. mirabilis S1959 and R45 cells. The saponins enhanced sensitivities of S and R P. mirabilis cells towards colistin and amipicillin. One may proposed that saponins binds to lipid A part of LPS may resulted on an increase in bacterial cell wall outer-membrane permeabilities and by that facilitated antibiotics penetration into the bacterial cells. In conclusion, the laser interferometric method is a useful tool for studies of lipopolysaccharide–antibiotic interactions even if the tested substances are not fully soluble in water.