ABSTRACT
The membrane fluidity of reduced-amphotericin B (AmB)-sensitivity Kluyveromyces lactis mutant strain is higher than that of the wild-type K. lactis strain. After culture of the K. lactis and K. lactis mutant cells in the presence of subinhibitory doses of AmB (10 and 125 mg/liter, respectively), the plasma membranes of both yeast strains also showed a higher fluidity than did those of control cells. High membrane fluidity was associated with changes in the structural properties of the membranes. Culture of the K. lactis and K. lactis mutant cells in the presence of AmB induced changes in membrane lipid contents. In particular, phospholipid contents were increased in both strains treated with AmB, compared with their corresponding counterparts. As a result, the sterol/phospholipid ratio decreased. The relative proportion of monounsaturated fatty acids also increased after AmB treatment. The saturated fatty acid/monounsaturated fatty acid ratio decreased in K. lactis and K. lactis mutant cells treated with AmB but also in K. lactis mutant control cells compared to that in the K. lactis wild strain. These changes in lipid composition explain the higher fluidity, which could represent a process of metabolic resistance of the yeasts to AmB.
Subject(s)
Amphotericin B/pharmacology , Antifungal Agents/pharmacology , Kluyveromyces/drug effects , Membrane Fluidity/drug effects , Buffers , Drug Resistance, Microbial/physiology , Kinetics , Kluyveromyces/physiology , Membrane Lipids/analysis , Osmolar Concentration , Spheroplasts/drug effects , Spheroplasts/physiology , Time FactorsABSTRACT
In Kluyveromyces lactis, the cell wall compositions of Kl (ATCC 96897), a wild sensitive strain, and Klm (ATCC 96896), a strain resistant to amphotericin B (AmB), were shown to be very different, since the walls in the latter were significantly enriched in hexosamine, but had a reduced content in phosphate and amino acid. In both strains, the cell walls limited their sensitivity to this antifungal agent. The absence of cell wall increased the sensitivity of the cells to this polyene by 5 to 10-fold. When the cells were treated with enzymes such as pronase and chitinase in order to change the cell wall structure just before inoculation, the yeasts appeared more resistant to the antibiotic. However, treatments with chymopapain and phospholipase C did not significantly change the sensitivity of the two strains to this agent. Cells treated with acid phosphatase displayed a longer lag phase than the control cells. In addition, when cultured in the presence of AmB, the cells were less sensitive to this agent. The present results reveal that both a change in the ionic charges of the cell wall and an alteration in the cell wall structure modified the sensitivity of these yeast strains to AmB.
