Twelve small plasmids isolated from avian strains of Pasteurella multocida were examined by restriction enzyme mapping, cross-hybridization, and minicell analysis. These plasmids contained sites for several commonly used restriction enzymes and ranged in size from 3.4 to 3.8 kilobases. Restriction enzyme maps of the 12 plasmids were similar and divided the plasmids into 3 families, designated pFS1, pFS2, and pFS4. Restriction fragments of pFS1 DNA isolated from strain X-73 were used to probe AvaI/HindIII/EcoRV digests of pFS2 and pFS4 DNA. The results of these hybridization experiments demonstrated that the plasmids found in all three families shared extensive regions of homology and may have originated from a common ancestor. Escherichia coli minicells containing recombinant plasmid constructs bearing fragments of pFS1 expressed two pFS1-specific peptides, 12.5 and 28 kilodaltons in size, suggesting that some P. multocida plasmid-encoded proteins can be expressed in E. coli. These results indicate that pFS may be useful as a genetic tool for moving DNA into and out of P. multocida, since it is small, contains common restriction sites, and encodes at least two genes that are recognized and expressed in E. coli.