Membrane fusion in exocytosis, intracellular trafficking, and enveloped viral infection is thought to be mediated by specialized proteins acting to merge membrane lipid bilayers. We now show that one class of naturally-occurring phospholipids, lysolipids, inhibits fusion between cell membranes, organelles, and between organelles and plasma membrane. Inhibition was reversible, did not correlate with lysis, and could be attributed to the molecular shape of lysolipids rather than to any specific chemical moiety. Fusion was arrested at a stage preceding fusion pore formation. Our results are consistent with the hypothesis that biological fusion, irrespective of trigger, involves the formation of a highly bent intermediate between membranes, the fusion stalk.