Two strains of Staphylococcus aureus (Newman and Tazaki) and their derived L-forms were cultured in serum-containing broth and the differences in their lipid compositions were analyzed. Cardiolipin accounted for more than 50% of the total phospholipid phosphorus in L-forms, but for less than 25% in parent bacteria. The cardiolipin content of L-forms was very high through all growth phases, although it increased gradually as growth proceeded. Significant amounts of cholesterol and its esters were present in parent strains and L-forms, all of which incorporated serum cholesterol into the cell membrane. On the other hand, they could be detected in the L-forms but not in the parent strains when they were cultured in serum-free broth. To examine the ability of L-forms to synthesize cholesterol, the cholesterol content of L-forms cultured in serum-free broth was compared with that of the medium. The results indicated that staphylococcal Lforms could synthesize cholesterol and its esters. These differences in lipid composition suggested that modification of membrane lipids may occur as an adaptational change in response to the disappearance of the cell wall.Bacterial L-forms which can be grown under appropriate conditions lack cell walls but have many of the biological characteristics of parent bacteria. The plasma membrane, as a result of being directly exposed to the external millieu, might be expected to undergo changes in both structure and function. Especially, stable L-forms which cannot revert to parent strains are more likely to acquire a membrane different from that of parent strains. Chemical analysis of stable L-form membrane, therefore, should provide information about the adaptational changes in the cell surface of a microorganism. There have been few reports of differences in membrane components between parent bacteria and derived L-forms (2, 19,26). Much of the attention has been focused, instead, on the presence or absence of cell wall substances in L-forms (16, 20,24) and the mode of L-form proliferation (5, 18). The authors have made a study on the differences in membrane components between Staphylococci and staphylococcal L-forms.The present work examines the phospholipid and neutral lipid composition of two stable L-forms of S. aureus and makes a comparison of these with those of the 435

The sterol which was present in two strains of a stable staphylococcal L-form was analyzed by gas-liquid chromatography and combined gas-liquid chromatography-mass spectrometry. The retention time of the sterol on gas-liquid chromatography was the same as that of authentic cholesterol. Analysis of the sterol by mass spectrometry showed a molecular ion at an m/e of 386 and the same patters of major ions above an m/e of 145 as those of authentic cholesterol. As a result, the sterol in staphylococcal L-form was identified as cholesterol. A parent strain and its L-form were cultured in medium containing [14C]acetate, and the