Physiological differences between cyclopropane fatty acid-deficient mutants and the parent strain of Streptococcus faecalis

Abstract

Cyclopropane fatty acids are a major fatty acid component of Streptococcus faecalis (ATCC 8043). The effect on the cell of the presence or absence of cyclopropanes is unknown. Comparative studies were made using mutants of S. faecalis which lacked cyclopropane fatty acids and the parent strain (SF/+), which contained cyclopropanes in normal quantities. Thin-layer chromatography studies confirmed that the cyclopropanes were associated with the cellular phospholipids. Factors which coincided with the initiation of cyclopropane fatty acid synthesis were investigated. The decline in medium pH as the culture grew coincided with the increased synthesis of cyclopropanes. When the normal decline in pH was retarded by growth in a more highly buffered medium, the synthesis of cyclopropanes still correlated with the medium pH and not with culture turbidity or nutritional changes in the medium. Growth studies were made using a rich medium which contained optimum amounts of thymidine and folic acid, and which contained no amethopterin. The maximum turbidities of the amethopterin-resistant strains and SF/ + were determined. The ratio of the maximum turbidity of SF/+ divided by the average maximum turbidity of the amethopterin-resistant mutants was 1.37 for normal growth at 37°C. For growth at 47°C, with 3.5% NaCl at 37 °C, with 3.5% NaCl at 47°C, or with 200 μg of sodium deoxycholate per ml, the ratio of growth turbidities was 6.42, 1.76, 9.70 and 6.75, respectively. The viability of SF/+ and the amethopterin-resistant strains was studied in a pH 4.0 sodium lactate medium. After 8 h exposure at 37 °C, at least 84% of the cyclopropane-deficient cells had lost viability, whereas SF/+ lost only 15%. In sodium lactate at pH 7.2 there was much less loss in viability with the mutants. NaCl at 47°C was shown to have a more deleterious effect on the amethopterin-resistant strains than on SF/+. In viability studies, over 95% of the cyclopropane-deficient cells died after 8 h exposure to 3.5% NaCl at 47°C, while SF/+ lost 10% over the same period. Uptake and/or retention of Na 2 35SO 4 and NaH 2 32PO 4 in the presence of 3.5% NaCl at 47°C was also lower in the amethopterin-resistant mutants than in SF/+. Viability studies showed that the mutant strains were much more susceptible to 400μg of sodium deoxycholate per ml than was the parent strain. Over 99% of the cyclopropane-deficient cells were dead after a 4-h exposure at 37°C, while 49% of the SF/+ cells retained viability under these conditions. These differences in susceptibility indicate that there may be a membrane-related physiological difference between SF/+ and the amethopterin-resistant mutants as a result of their cyclopropane fatty acid deficiency.