Feeding trials using purified test diets were conducted in order to clarify the requirements of essential fatty acids (EFA) for the larval stages of an Ayu Plecoglossus altivelis. The weight gains were low in the larvae fed on the lipid-free and ω3 fatty acid-deficient diets. The supplement of linolenic acid (18:3ω3) or eicosapentaenoic acid (20:5ω3) to the diet containing 8% levels of lipids, oleic acid (18:1ω9), or egg lecithin-18:1ω9 (3:5, w/w) improved the weight gain of the Ayu larvae, whereas that of linoleic acid (18:2ω6) enhanced growth very little. The growthpromoting effect of 18:3ω3 was almost equal to that of 20:5ω3. These results indicated that the larval Ayu requires 18:3ω3 and 20:5ω3 as EFA for growth.