FACTORS AFFECTING THE PERMEABILITY OF PSEUDOMONAS-AERUGINOSA CELL-WALLS TOWARD LIPOPHILIC COMPOUNDS - EFFECTS OF ULTRASOUND AND CELL AGE

The objective of this research was to elucidate the factors effecting the permeability of cell membranes of gram-negative bacteria toward hy drophobic compounds, Ultrasound treatment, cell age, and, the phase st ate of phospholipid membranes were considered, Spin-labeling EPR metho d was used to quantify the penetration and distribution of a lipophili c spin probe, 16-dosylstearic acid (16-DS), in Pseudomonas aeruginosa cell membranes, This bacterium was chosen because of its reported resi stance to the action of hydrophobic antibiotics caused by the low perm eability of the outer cell membrane for hydrophobic compounds, EPR spe ctra were collected from cell pellets and cell lysates, The overall sp ill probe uptake was measured in 10% SDS-cell lysates, Lysis with 0.6% SDS revealed the fraction of the probe located in membrane sites read ily accessible to the surfactant. The results indicated a structural h eterogeneity of aeruginosa membranes, with the presence of structurall y ''stronger'' and ''weaker'' sites characterized by different suscept ibility to the SDS treatment, The intracellular concentration of 16-DS was higher in insonated cells and increased linearly with the sonicat ion power. EPR spectra indicated that ultrasound enhanced the penetrat ion of the probe into the structurally stronger sites of the inner and outer cell membranes, The effect of ultrasound on the cell membranes was transient in that the initial membrane permeability was restored u pon termination of the ultrasound treatment. These results suggest tha t the resistance of gram-negative bacteria to the action of hydrophobi c antibiotics was caused by a low permeability of the outer cell membr anes. This resistance may be reduced by the simultaneous application o f antibiotic and ultrasound. This hypothesis was confirmed in our expe riments with P. aeruginosa exposed to erythromycin. (C) 1997 Academic Press.