The composition of phospholipids and their fatty acids was investigated in the infective juveniles (IJs) of four species of entomopathogenic nematodes: Steinernema carpocapsae (strain All), S. riobravis (strain Biosys 355), S. feltiae (strain UK76) and S. glaseri (strain NC). In newly emerged IJs of the four species, phospholipids comprised 15–18% dry weight of the total lipids (or 5–6% dry weight of the nematode), and phosphatidylethanolamine and phosphatidylcholine constituted about 40 and 30%, respectively, of the total phospholipids, with phosphatidylserine (PS) and phosphatidylinositol (PI) collectively accounting for about 25%. Qualitatively, the four species had identical total phospholipid (combined non-acidic and acidic fractions) fatty acid profiles, although there were some differences in the relative proportions (mol%) of specific fatty acids. The fatty acid composition of the total phospholipids in newly emerged IJs was dominated by C16 fatty acids, specifically C16:0 (14–18%), C16:1n-7 (up to 20%) and C16:4 (up to 26%), whereas the major C18 fatty acid was C18:1n-9 (20–23%). Polyunsaturated C20 fatty acids collectively made up 8–13% of the total composition. When newly emerged IJs were stored in distilled water at 25°C, the proportions of C16:0 and C16:4 decreased with storage time, whereas C16:3n-3 increased (by 30-fold in S. glaseri). These changes were mostly observed in the acidic phospholipid fraction (mainly PI and PS). No evidence was found for a correlation between the fatty acid composition of the phospholipids and the relative ability of the IJs of the four Steinernema species to survive desiccation stress.