A reduction in the dietary concentration of choline, an essential nutrient for Drosophila melanogaster, from the optimal concentration of 80 micrograms/ml of defined medium to 8 micrograms/ml diminished the level of tissue phosphatidylcholine to less than one-third the normal level in third instar larvae without significantly altering the amount of phosphatidylethanolamine. 2. The rates of synthesis of phospholipids, triglycerides, diglycerides and monoglycerides were reduced by the choline-deficiency, and the chain length of fatty acids in lipids was shortened. 3. The activity of succinic dehydrogenase, a mitochondrial enzyme, was decreased by the deficiency, but the activities of fumarase, sn-glycerol-3-phosphate dehydrogenase, alcohol dehydrogenase, sn-glycerol-3-phosphate oxidase and fatty acid synthetase were unaffected. A choline-deficiency did not alter the ultrastructure of mitochondria of larval fat body cells. 4. Choline-deficient individuals were more susceptible to the toxic effects of ethanol during larval and pupal development, and less adept at utilizing ethanol as a substrate for adult tissue synthesis.