Amber disease, a chronic disease of Costelytra zealandica (Coleoptera: Scarabaeidae) larvae, is caused by some isolates of the bacteria, Serratia entomophila and Serratia proteamaculans (Enterobacteriaceae). A large plasmid was visualized in some isolates of these two Serratia species. Further investigation showed that 18 of 19 pathogenic isolates of Serratia spp. were found to have a megaplasmid, estimated to be 105 kb. The one virulent isolate lacking a 105-kb plasmid had two smaller plasmids. The 105-kd plasmid was present in only 1 of 22 naturally occurring, nonpathogenic isolates examined. Plasmid DNA isolated from pathogenic isolates of each species showed identical restriction patterns. Heat-treated derivatives of pathogenic S. entomophila and S. proteamaculans isolates, which lost disease-causing ability, also lost the 105-kb plasmid. Among isolates of 7 other species of Serratia, single isolates of Serratia grimesii and Serratia ficaria were found to have a similarly sized plasmid, but caused no disease symptoms. The implications of the association between the plasmid and amber disease are discussed.